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Sample records for orthogonal trna-aminoacyl-trna synthetase

  1. Methods and composition for the production of orthogonal tRNA-aminoacyltRNA synthetase pairs

    DOEpatents

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Anderson, John Christopher [San Diego, CA; Chin, Jason [Cambridge, GB; Liu, David R [Lexington, MA; Magliery, Thomas J [North Haven, CT; Meggers, Eric L [Philadelphia, PA; Mehl, Ryan Aaron [Lancaster, PA; Pastrnak, Miro [San Diego, CA; Santoro, Steven William [Cambridge, MA; Zhang, Zhiwen [San Diego, CA

    2008-04-08

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  2. Methods and composition for the production of orthogonal tRNA-aminoacyltRNA synthetase pairs

    DOEpatents

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Anderson, John Christopher [San Diego, CA; Chin, Jason [Cambridge, GB; Liu, David R [Lexington, MA; Magliery, Thomas J [North Haven, CT; Meggers, Eric L [Philadelphia, PA; Mehl, Ryan Aaron [Lancaster, PA; Pastrnak, Miro [San Diego, CA; Santoro, Steven William [Cambridge, MA; Zhang, Zhiwen [San Diego, CA

    2012-05-22

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  3. Methods and compositions for the production of orthogonal tRNA-aminoacyl-tRNA synthetase pairs

    DOEpatents

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Anderson, John Christopher [San Diego, CA; Chin, Jason W [San Diego, CA; Liu, David R [Lexington, MA; Magliery, Thomas J [North Haven, CT; Meggers, Eric L [Philadelphia, PA; Mehl, Ryan Aaron [San Diego, CA; Pastrnak, Miro [San Diego, CA; Santoro, Stephen William [San Diego, CA; Zhang, Zhiwen [San Diego, CA

    2011-09-06

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  4. Methods and compositions for the production of orthogonal tRNA-aminoacyl tRNA synthetase pairs

    DOEpatents

    Schultz, Peter G.; Wang, Lei; Anderson, John Christopher; Chin, Jason W.; Liu, David R.; Magliery, Thomas J.; Meggers, Eric L.; Mehl, Ryan Aaron; Pastrnak, Miro; Santoro, Stephen William; Zhang, Zhiwen

    2015-10-20

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  5. Methods and composition for the production of orthogonal tRNA-aminoacyl tRNA synthetase pairs

    DOEpatents

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Anderson, John Christopher [San Diego, CA; Chin, Jason W [San Diego, CA; Liu, David R [Lexington, MA; Magliery, Thomas J [North Haven, CT; Meggers, Eric L [Philadelphia, PA; Mehl, Ryan Aaron [San Diego, CA; Pastrnak, Miro [San Diego, CA; Santoro, Stephen William [San Diego, CA; Zhang, Zhiwen [San Diego, CA

    2012-05-08

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  6. Methods and compositions for the production of orthogonal tRNA-aminoacyl tRNA synthetase pairs

    SciTech Connect

    Schultz, Peter; Wang, Lei; Anderson, John Christopher; Chin, Jason; Liu, David R.; Magliery, Thomas J.; Meggers, Eric L.; Mehl, Ryan Aaron; Pastrnak, Miro; Santoro, Stephen William; Zhang, Zhiwen

    2006-08-01

    This invention provides compositions and methods for generating components of protein biosynthetic machinery including orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases. Methods for identifying orthogonal pairs are also provided. These components can be used to incorporate unnatural amino acids into proteins in vivo.

  7. Compositions of orthogonal lysyl-tRNA and aminoacyl-tRNA synthetase pairs and uses thereof

    DOEpatents

    Anderson, J. Christopher; Wu, Ning; Santoro, Stephen; Schultz, Peter G

    2014-03-11

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal lysyl-tRNAs, orthogonal lysyl-aminoacyl-tRNA synthetases, and orthogonal pairs of lysyl-tRNAs/synthetases, which incorporate homoglutamines into proteins are provided in response to a four base codon. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with homoglutamines using these orthogonal pairs.

  8. Study of the Binding Energies between Unnatural Amino Acids and Engineered Orthogonal Tyrosyl-tRNA Synthetases

    NASA Astrophysics Data System (ADS)

    Ren, Wei; Truong, Tan M.; Ai, Hui-Wang

    2015-07-01

    We utilized several computational approaches to evaluate the binding energies of tyrosine (Tyr) and several unnatural Tyr analogs, to several orthogonal aaRSes derived from Methanocaldococcus jannaschii and Escherichia coli tyrosyl-tRNA synthetases. The present study reveals the following: (1) AutoDock Vina and ROSETTA were able to distinguish binding energy differences for individual pairs of favorable and unfavorable aaRS-amino acid complexes, but were unable to cluster together all experimentally verified favorable complexes from unfavorable aaRS-Tyr complexes; (2) MD-MM/PBSA provided the best prediction accuracy in terms of clustering favorable and unfavorable enzyme-substrate complexes, but also required the highest computational cost; and (3) MM/PBSA based on single energy-minimized structures has a significantly lower computational cost compared to MD-MM/PBSA, but still produced sufficiently accurate predictions to cluster aaRS-amino acid interactions. Although amino acid-aaRS binding is just the first step in a complex series of processes to acylate a tRNA with its corresponding amino acid, the difference in binding energy, as shown by MD-MM/PBSA, is important for a mutant orthogonal aaRS to distinguish between a favorable unnatural amino acid (unAA) substrate from unfavorable natural amino acid substrates. Our computational study should assist further designing and engineering of orthogonal aaRSes for the genetic encoding of novel unAAs.

  9. Study of the Binding Energies between Unnatural Amino Acids and Engineered Orthogonal Tyrosyl-tRNA Synthetases

    PubMed Central

    Ren, Wei; Truong, Tan M.; Ai, Hui-wang

    2015-01-01

    We utilized several computational approaches to evaluate the binding energies of tyrosine (Tyr) and several unnatural Tyr analogs, to several orthogonal aaRSes derived from Methanocaldococcus jannaschii and Escherichia coli tyrosyl-tRNA synthetases. The present study reveals the following: (1) AutoDock Vina and ROSETTA were able to distinguish binding energy differences for individual pairs of favorable and unfavorable aaRS-amino acid complexes, but were unable to cluster together all experimentally verified favorable complexes from unfavorable aaRS-Tyr complexes; (2) MD-MM/PBSA provided the best prediction accuracy in terms of clustering favorable and unfavorable enzyme-substrate complexes, but also required the highest computational cost; and (3) MM/PBSA based on single energy-minimized structures has a significantly lower computational cost compared to MD-MM/PBSA, but still produced sufficiently accurate predictions to cluster aaRS-amino acid interactions. Although amino acid-aaRS binding is just the first step in a complex series of processes to acylate a tRNA with its corresponding amino acid, the difference in binding energy, as shown by MD-MM/PBSA, is important for a mutant orthogonal aaRS to distinguish between a favorable unnatural amino acid (unAA) substrate from unfavorable natural amino acid substrates. Our computational study should assist further designing and engineering of orthogonal aaRSes for the genetic encoding of novel unAAs. PMID:26220470

  10. Identification of potent inhibitors of the Trypanosoma brucei methionyl-tRNA synthetase via high-throughput orthogonal screening.

    PubMed

    Pedró-Rosa, Laura; Buckner, Frederick S; Ranade, Ranae M; Eberhart, Christina; Madoux, Franck; Gillespie, J Robert; Koh, Cho Yeow; Brown, Steven; Lohse, Jacqueline; Verlinde, Christophe L M; Fan, Erkang; Bannister, Thomas; Scampavia, Louis; Hol, Wim G J; Spicer, Timothy; Hodder, Peter

    2015-01-01

    Improved therapies for the treatment of Trypanosoma brucei, the etiological agent of the neglected tropical disease human African trypanosomiasis, are urgently needed. We targeted T. brucei methionyl-tRNA synthetase (MetRS), an aminoacyl-tRNA synthase (aaRS), which is considered an important drug target due to its role in protein synthesis, cell survival, and its significant differences in structure from its mammalian ortholog. Previous work using RNA interference of MetRS demonstrated growth inhibition of T. brucei, further validating it as an attractive target. We report the development and implementation of two orthogonal high-throughput screening assays to identify inhibitors of T. brucei MetRS. First, a chemiluminescence assay was implemented in a 1536-well plate format and used to monitor adenosine triphosphate depletion during the aminoacylation reaction. Hit confirmation then used a counterscreen in which adenosine monophosphate production was assessed using fluorescence polarization technology. In addition, a miniaturized cell viability assay was used to triage cytotoxic compounds. Finally, lower throughput assays involving whole parasite growth inhibition of both human and parasite MetRS were used to analyze compound selectivity and efficacy. The outcome of this high-throughput screening campaign has led to the discovery of 19 potent and selective T. brucei MetRS inhibitors.

  11. Orthogonal translation components for the in vivo incorporation of unnatural amino acids

    DOEpatents

    Schultz, Peter G.; Alfonta, Lital; Chittuluru, Johnathan R.; Deiters, Alexander; Groff, Dan; Summerer, Daniel; Tsao, Meng -Lin; Wang, Jiangyun; Wu, Ning; Xie, Jianming; Zeng, Huaqiang; Seyedsayamdost, Mohammad; Turner, James

    2015-08-11

    The invention relates to orthogonal pairs of tRNAs and aminoacyl-tRNA synthetase that can incorporate unnatural amino acid into proteins produced in eubacterial host cells such as E. coli, or in a eukaryotic host such as a yeast cell. The invention provides, for example but not limited to, novel orthogonal synthetases, methods for identifying and making the novel synthetases, methods for producing proteins containing unnatural amino acids, and translation systems.

  12. Orthogonal translation components for the in vivo incorporation of unnatural amino acids

    SciTech Connect

    Schultz, Peter G.; Xie, Jianming; Zeng, Huaqiang

    2012-07-10

    The invention relates to orthogonal pairs of tRNAs and aminoacyl-tRNA synthetases that can incorporate unnatural amino acids into proteins produced in eubacterial host cells such as E. coli, or in a eukaryotic host such as a yeast cell. The invention provides, for example but not limited to, novel orthogonal synthetases, methods for identifying and making the novel synthetases, methods for producing proteins containing unnatural amino acids, and translation systems.

  13. [Anti-synthetase syndrome].

    PubMed

    Novak, Srdan

    2012-01-01

    Antysynthetase syndrome is considered as a group ofidiopathic inflammatory myositis with charcteristic serologic hallmark--antibodies which recognise the aminoacyl-tRNA synthetasses (ARS). Clinical picture of those patients contains myositis and/or intersticial lung disease (ILD) and/or arthritis and/or fever and/or Raynaud phenomenon and sometimes characteristic look of mechanic's hands. Myositis can be overt, sometimes even absent, while IBP is major cause of morbidity and determines the outcome of the disease. Untill now eight different any-synthetase autoantibodies are recognised, and most frequent are findings of anti-histidyl-tRNa synthetase antibodies. Patients with other ARS autoantibodies usually have severe ILD. Drug of choice are steroids in dosage of 1 mg/kg with immunosupresive agent (azatioprin or methotrexate) while in severe IBP cyclophosphamide is needed. Recently succsesful treatment with rituximab in combination with cyclophosphamide is reported.

  14. Pyrrolysyl-tRNA synthetase, an aminoacyl-tRNA synthetase for genetic code expansion

    PubMed Central

    Crnković, Ana; Suzuki, Tateki; Söll, Dieter; Reynolds, Noah M.

    2016-01-01

    Genetic code expansion (GCE) has become a central topic of synthetic biology. GCE relies on engineered aminoacyl-tRNA synthetases (aaRSs) and a cognate tRNA species to allow codon reassignment by co-translational insertion of non-canonical amino acids (ncAAs) into proteins. Introduction of such amino acids increases the chemical diversity of recombinant proteins endowing them with novel properties. Such proteins serve in sophisticated biochemical and biophysical studies both in vitro and in vivo, they may become unique biomaterials or therapeutic agents, and they afford metabolic dependence of genetically modified organisms for biocontainment purposes. In the Methanosarcinaceae the incorporation of the 22nd genetically encoded amino acid, pyrrolysine (Pyl), is facilitated by pyrrolysyl-tRNA synthetase (PylRS) and the cognate UAG-recognizing tRNAPyl. This unique aaRS•tRNA pair functions as an orthogonal translation system (OTS) in most model organisms. The facile directed evolution of the large PylRS active site to accommodate many ncAAs, and the enzyme’s anticodon-blind specific recognition of the cognate tRNAPyl make this system highly amenable for GCE purposes. The remarkable polyspecificity of PylRS has been exploited to incorporate >100 different ncAAs into proteins. Here we review the Pyl-OT system and selected GCE applications to examine the properties of an effective OTS. PMID:28239189

  15. Orthogonal transform feasibility study

    NASA Technical Reports Server (NTRS)

    Robinson, G. S.

    1971-01-01

    The application of various orthogonal transformations to communication was investigated, with particular emphasis placed on speech and visual signal processing. The fundamentals of the one- and two-dimensional orthogonal transforms and their application to speech and visual signals are treated in detail.

  16. Genetics Home Reference: glutathione synthetase deficiency

    MedlinePlus

    ... Facebook Share on Twitter Your Guide to Understanding Genetic Conditions Search MENU Toggle navigation Home Page Search ... Conditions Genes Chromosomes & mtDNA Resources Help Me Understand Genetics Home Health Conditions glutathione synthetase deficiency glutathione synthetase ...

  17. Coherent orthogonal polynomials

    SciTech Connect

    Celeghini, E.; Olmo, M.A. del

    2013-08-15

    We discuss a fundamental characteristic of orthogonal polynomials, like the existence of a Lie algebra behind them, which can be added to their other relevant aspects. At the basis of the complete framework for orthogonal polynomials we include thus–in addition to differential equations, recurrence relations, Hilbert spaces and square integrable functions–Lie algebra theory. We start here from the square integrable functions on the open connected subset of the real line whose bases are related to orthogonal polynomials. All these one-dimensional continuous spaces allow, besides the standard uncountable basis (|x〉), for an alternative countable basis (|n〉). The matrix elements that relate these two bases are essentially the orthogonal polynomials: Hermite polynomials for the line and Laguerre and Legendre polynomials for the half-line and the line interval, respectively. Differential recurrence relations of orthogonal polynomials allow us to realize that they determine an infinite-dimensional irreducible representation of a non-compact Lie algebra, whose second order Casimir C gives rise to the second order differential equation that defines the corresponding family of orthogonal polynomials. Thus, the Weyl–Heisenberg algebra h(1) with C=0 for Hermite polynomials and su(1,1) with C=−1/4 for Laguerre and Legendre polynomials are obtained. Starting from the orthogonal polynomials the Lie algebra is extended both to the whole space of the L{sup 2} functions and to the corresponding Universal Enveloping Algebra and transformation group. Generalized coherent states from each vector in the space L{sup 2} and, in particular, generalized coherent polynomials are thus obtained. -- Highlights: •Fundamental characteristic of orthogonal polynomials (OP): existence of a Lie algebra. •Differential recurrence relations of OP determine a unitary representation of a non-compact Lie group. •2nd order Casimir originates a 2nd order differential equation that defines

  18. Rational design of an orthogonal tryptophanyl nonsense suppressor tRNA

    PubMed Central

    Hughes, Randall A.; Ellington, Andrew D.

    2010-01-01

    While a number of aminoacyl tRNA synthetase (aaRS):tRNA pairs have been engineered to alter or expand the genetic code, only the Methanococcus jannaschii tyrosyl tRNA synthetase and tRNA have been used extensively in bacteria, limiting the types and numbers of unnatural amino acids that can be utilized at any one time to expand the genetic code. In order to expand the number and type of aaRS/tRNA pairs available for engineering bacterial genetic codes, we have developed an orthogonal tryptophanyl tRNA synthetase and tRNA pair, derived from Saccharomyces cerevisiae. In the process of developing an amber suppressor tRNA, we discovered that the Escherichia coli lysyl tRNA synthetase was responsible for misacylating the initial amber suppressor version of the yeast tryptophanyl tRNA. It was discovered that modification of the G:C content of the anticodon stem and therefore reducing the structural flexibility of this stem eliminated misacylation by the E. coli lysyl tRNA synthetase, and led to the development of a functional, orthogonal suppressor pair that should prove useful for the incorporation of bulky, unnatural amino acids into the genetic code. Our results provide insight into the role of tRNA flexibility in molecular recognition and the engineering and evolution of tRNA specificity. PMID:20571084

  19. Hepatocytes explanted in the spleen preferentially express carbamoylphosphate synthetase rather than glutamine synthetase.

    PubMed

    Lamers, W H; Been, W; Charles, R; Moorman, A F

    1990-10-01

    Urea cycle enzymes and glutamine synthetase are essential for NH3 detoxification and systemic pH homeostasis in mammals. Carbamoylphosphate synthetase, the first and flux-determining enzyme of the cycle, is found only in a large periportal compartment, and glutamine synthetase is found only in a small, complementary pericentral compartment. Because it is not possible to manipulate experimentally the intrahepatic distribution of carbamoylphosphate synthetase and glutamine synthetase, we looked for conditions in which explanted hepatocytes would exhibit either the carbamoylphosphate synthetase phenotype or glutamine synthetase phenotype. In the spleen hepatocytes either settle as individual cells or in small agglomerates. The dispersed cells only express the carbamoylphosphate synthetase phenotype. Within the agglomerates, sinusoids that drain on venules develop. Hepatocytes surrounding the venules stain only weakly for carbamoylphosphate synthetase but are strongly positive for glutamine synthetase. These observations were made for explanted embryonic hepatocytes (no prior expression of either carbamoylphosphate synthetase or glutamine synthetase), neonatal hepatocytes (compartments of gene expression not yet established) and adult periportal and pericentral hepatocytes.

  20. Genetics Home Reference: carbamoyl phosphate synthetase I deficiency

    MedlinePlus

    ... Health Conditions carbamoyl phosphate synthetase I deficiency carbamoyl phosphate synthetase I deficiency Printable PDF Open All Close ... to view the expand/collapse boxes. Description Carbamoyl phosphate synthetase I deficiency is an inherited disorder that ...

  1. Radioimmune assay of human platelet prostaglandin synthetase

    SciTech Connect

    Roth, G.J.; Machuga, E.T.

    1982-02-01

    Normal platelet function depends, in part, on platelet PG synthesis. PG synthetase (cyclo-oxygenase) catalyzes the first step in PG synthesis, the formation of PGH/sub 2/ from arachidonic acid. Inhibition of the enzyme by ASA results in an abnormality in the platelet release reaction. Patients with pparent congenital abnormalities in the enzyme have been described, and the effects have been referred to as ''aspirin-like'' defects of the platelet function. These patients lack platelet PG synthetase activity, but the actual content of PG synthetase protein in these individuals' platelets is unknown. Therefore an RIA for human platelet PG synthetase would provide new information, useful in assessing the aspirin-like defects of platelet function. An RIA for human platelet PG synthetase is described. The assay utilizes a rabbit antibody directed against the enzyme and (/sup 125/I)-labelled sheep PG synthetase as antigen. The human platelet enzyme is assayed by its ability to inhibit precipitation of the (/sup 125/I)antigen. The assay is sensitive to 1 ng of enzyme. By the immune assay, human platelets contain approximately 1200 ng of PG synethetase protein per 1.5 mg of platelet protein (approximately 10/sup 9/ platelets). This content corresponds to 10,000 enzyme molecules per platelet. The assay provides a rapid and convenient assay for the human platelet enzyme, and it can be applied to the assessment of patients with apparent platelet PG synthetase (cyclo-oxygenase) deficiency.

  2. Gene encoding plant asparagine synthetase

    DOEpatents

    Coruzzi, Gloria M.; Tsai, Fong-Ying

    1993-10-26

    The identification and cloning of the gene(s) for plant asparagine synthetase (AS), an important enzyme involved in the formation of asparagine, a major nitrogen transport compound of higher plants is described. Expression vectors constructed with the AS coding sequence may be utilized to produce plant AS; to engineer herbicide resistant plants, salt/drought tolerant plants or pathogen resistant plants; as a dominant selectable marker; or to select for novel herbicides or compounds useful as agents that synchronize plant cells in culture. The promoter for plant AS, which directs high levels of gene expression and is induced in an organ specific manner and by darkness, is also described. The AS promoter may be used to direct the expression of heterologous coding sequences in appropriate hosts.

  3. Evolving Orthogonal Suppressor tRNAs To Incorporate Modified Amino Acids.

    PubMed

    Maranhao, Andre C; Ellington, Andrew D

    2017-01-20

    There have been considerable advancements in the incorporation of noncanonical amino acids (ncAA) into proteins over the last two decades. The most widely used method for site-specific incorporation of noncanonical amino acids, amber stop codon suppression, typically employs an orthogonal translation system (OTS) consisting of a heterologous aminoacyl-tRNA synthetase:tRNA pair that can potentially expand an organism's genetic code. However, the orthogonal machinery sometimes imposes fitness costs on an organism, in part due to mischarging and a lack of specificity. Using compartmentalized partnered replication (CPR) and a newly developed pheS negative selection, we evolved several new orthogonal Methanocaldococcus jannaschii (Mj) tRNA variants tRNAs with increased amber suppression activity, but that also showed up to 3-fold reduction in promiscuous aminoacylation by endogenous aminoacyl-tRNA synthetases (aaRSs). The increased orthogonality of these variants greatly reduced organismal fitness costs associated in part due to tRNA mischarging. Using these methods, we were also able to evolve tRNAs that supported the specific incorporation of 3-halo-tyrosines (3-Cl-Y, 3-Br-Y, and 3-I-Y) in E. coli.

  4. Genetically expanded cell-free protein synthesis using endogenous pyrrolysyl orthogonal translation system.

    PubMed

    Chemla, Yonatan; Ozer, Eden; Schlesinger, Orr; Noireaux, Vincent; Alfonta, Lital

    2015-08-01

    Cell-free protein synthesis offers a facile and rapid method for synthesizing, monitoring, analyzing, and purifying proteins from a DNA template. At the same time, genetic code expansion methods are gaining attention due to their ability to site-specifically incorporate unnatural amino acids (UAAs) into proteins via ribosomal translation. These systems are based on the exogenous addition of an orthogonal translation system (OTS), comprising an orthogonal tRNA, and orthogonal aminoacyl tRNA synthetase (aaRS), to the cell-free reaction mixture. However, these components are unstable and their preparation is labor-intensive, hence introducing a major challenge to the system. Here, we report on an approach that significantly reduces the complexity, effort and time needed to express UAA-containing proteins while increasing stability and realizing maximal suppression efficiency. We demonstrate an endogenously introduced orthogonal pair that enables the use of the valuable yet insoluble pyrrolysyl-tRNA synthetase in a cell-free system, thereby expanding the genetic repertoire that can be utilized in vitro and enabling new possibilities for bioengineering. With the high stability and efficiency of our system, we offer an improved and accessible platform for UAA incorporation into proteins.

  5. Orthogonal Regression: A Teaching Perspective

    ERIC Educational Resources Information Center

    Carr, James R.

    2012-01-01

    A well-known approach to linear least squares regression is that which involves minimizing the sum of squared orthogonal projections of data points onto the best fit line. This form of regression is known as orthogonal regression, and the linear model that it yields is known as the major axis. A similar method, reduced major axis regression, is…

  6. Orthogonal Regression: A Teaching Perspective

    ERIC Educational Resources Information Center

    Carr, James R.

    2012-01-01

    A well-known approach to linear least squares regression is that which involves minimizing the sum of squared orthogonal projections of data points onto the best fit line. This form of regression is known as orthogonal regression, and the linear model that it yields is known as the major axis. A similar method, reduced major axis regression, is…

  7. Genetics Home Reference: holocarboxylase synthetase deficiency

    MedlinePlus

    ... use of biotin, a B vitamin found in foods such as liver, egg yolks, and milk. Holocarboxylase synthetase attaches biotin to certain enzymes that are essential for the normal production and breakdown of proteins, fats, and carbohydrates in ...

  8. "Orthogonality" in Learning and Assessment

    ERIC Educational Resources Information Center

    Leslie, David

    2014-01-01

    This chapter proposes a simple framework, "orthogonality," to help clarify what stakeholders think about learning in college, how we assess outcomes, and how clear assessment methods might help increase confidence in returns on investment.

  9. Orthogonal polynomials and deformed oscillators

    NASA Astrophysics Data System (ADS)

    Borzov, V. V.; Damaskinsky, E. V.

    2015-10-01

    In the example of the Fibonacci oscillator, we discuss the construction of oscillator-like systems associated with orthogonal polynomials. We also consider the question of the dimensions of the corresponding Lie algebras.

  10. Compact orthogonal NMR field sensor

    DOEpatents

    Gerald, II, Rex E.; Rathke, Jerome W.

    2009-02-03

    A Compact Orthogonal Field Sensor for emitting two orthogonal electro-magnetic fields in a common space. More particularly, a replacement inductor for existing NMR (Nuclear Magnetic Resonance) sensors to allow for NMR imaging. The Compact Orthogonal Field Sensor has a conductive coil and a central conductor electrically connected in series. The central conductor is at least partially surrounded by the coil. The coil and central conductor are electrically or electro-magnetically connected to a device having a means for producing or inducing a current through the coil and central conductor. The Compact Orthogonal Field Sensor can be used in NMR imaging applications to determine the position and the associated NMR spectrum of a sample within the electro-magnetic field of the central conductor.

  11. "Orthogonality" in Learning and Assessment

    ERIC Educational Resources Information Center

    Leslie, David

    2014-01-01

    This chapter proposes a simple framework, "orthogonality," to help clarify what stakeholders think about learning in college, how we assess outcomes, and how clear assessment methods might help increase confidence in returns on investment.

  12. Generations of orthogonal surface coordinates

    NASA Technical Reports Server (NTRS)

    Blottner, F. G.; Moreno, J. B.

    1980-01-01

    Two generation methods were developed for three dimensional flows where the computational domain normal to the surface is small. With this restriction the coordinate system requires orthogonality only at the body surface. The first method uses the orthogonal condition in finite-difference form to determine the surface coordinates with the metric coefficients and curvature of the coordinate lines calculated numerically. The second method obtains analytical expressions for the metric coefficients and for the curvature of the coordinate lines.

  13. Non-Orthogonal Iris Segmentation

    DTIC Science & Technology

    2005-05-09

    pixels ( eyelashes , eyelids, etc.). For example, if all three quality bounds are close in proximity, the iris pattern that was segmented...iris pattern that was segmented by the algorithm contains eyelashes and additional extraneous information. If all automatically generated “test” mask...orthogonal iris images (see Figs. 25-28). This is primarily explained by the increased presence of eyelashes in the non-orthogonal iris masks. As the angle

  14. Fatty Acid Synthetase of Saccharomyces cerevisiae

    PubMed Central

    Klein, Harold P.; Volkmann, Carol M.; Chao, Fu-Chuan

    1967-01-01

    A light particle fraction of Saccharomyces cerevisiae, obtained from the crude ribosomal material, and containing the fatty acid synthetase, consisted primarily of 27S and 47S components. This fraction has a protein-ribonucleic acid ratio of about 13. Electron micrographs showed particles ranging in diameter between 100 and 300 A in this material. By use of density gradient analysis, the fatty acid synthetase was found in the 47S component. This component contained particles which were predominantly 300 A in diameter and which were considerably flatter than ribosomes, and it consisted almost entirely of protein. Images PMID:6025308

  15. Orthogonal-polarization multipulse sequences

    NASA Astrophysics Data System (ADS)

    Grydeland, T.; Gustavsson, B.

    2011-02-01

    It is well known that using orthogonal polarizations for subpulses in multipulse sequences can be used to reduce clutter contributions in these modes. In this paper we show that further improvements are achieved if the orthogonality is taken into account already when constructing the codes. Using orthogonal polarizations, one can use denser transmission patterns, including elementary pulses without gaps between them, patterns that have severe range ambiguities when only a single polarization is used. Furthermore, correlations are computed separately for each combination of elementary pulse polarizations. Consequently, it is possible to have longer multipulse sequences without gaps in the lag sequence, it is possible to compute the odd lags as well as the even ones, and it is permissible to have some lags multiply obtained without range ambiguity. This means that using orthogonal polarizations when creating the multipulse transmission pattern gives flexibility well beyond the single-polarization case. This flexibility can be used to design patterns suited to particular experimental requirements. Furthermore, we point out that the improvement in clutter might have a more dramatic impact than is generally realized, particularly in high-SNR situations where the improvement in speed is up to a factor of 4. Examples are given of single- and multiple-frequency patterns that are not usable if only one polarization is available. Although all incoherent scatter radars in use today, except Jicamarca, lack orthogonal polarization capabilities, designers of the next generation of radars might find the improvements described herein to be of interest.

  16. Orthogonality of spherical harmonic coefficients

    NASA Technical Reports Server (NTRS)

    Mcleod, M. G.

    1980-01-01

    Orthogonality relations are obtained for the spherical harmonic coefficients of functions defined on the surface of a sphere. Following a brief discussion of the orthogonality of Fourier series coefficients, consideration is given to the values averaged over all orientations of the coordinate system of the spherical harmonic coefficients of a function defined on the surface of a sphere that can be expressed in terms of Legendre polynomials for the special case where the function is the sum of two delta functions located at two different points on the sphere, and for the case of an essentially arbitrary function. It is noted that the orthogonality relations derived have found applications in statistical studies of the geomagnetic field.

  17. Orthogonality of spherical harmonic coefficients

    NASA Technical Reports Server (NTRS)

    Mcleod, M. G.

    1980-01-01

    Orthogonality relations are obtained for the spherical harmonic coefficients of functions defined on the surface of a sphere. Following a brief discussion of the orthogonality of Fourier series coefficients, consideration is given to the values averaged over all orientations of the coordinate system of the spherical harmonic coefficients of a function defined on the surface of a sphere that can be expressed in terms of Legendre polynomials for the special case where the function is the sum of two delta functions located at two different points on the sphere, and for the case of an essentially arbitrary function. It is noted that the orthogonality relations derived have found applications in statistical studies of the geomagnetic field.

  18. A euryarchaeal lysyl-tRNA synthetase: resemblance to class I synthetases.

    PubMed

    Ibba, M; Morgan, S; Curnow, A W; Pridmore, D R; Vothknecht, U C; Gardner, W; Lin, W; Woese, C R; Söll, D

    1997-11-07

    The sequencing of euryarchaeal genomes has suggested that the essential protein lysyl-transfer RNA (tRNA) synthetase (LysRS) is absent from such organisms. However, a single 62-kilodalton protein with canonical LysRS activity was purified from Methanococcus maripaludis, and the gene that encodes this protein was cloned. The predicted amino acid sequence of M. maripaludis LysRS is similar to open reading frames of unassigned function in both Methanobacterium thermoautotrophicum and Methanococcus jannaschii but is unrelated to canonical LysRS proteins reported in eubacteria, eukaryotes, and the crenarchaeote Sulfolobus solfataricus. The presence of amino acid motifs characteristic of the Rossmann dinucleotide-binding domain identifies M. maripaludis LysRS as a class I aminoacyl-tRNA synthetase, in contrast to the known examples of this enzyme, which are class II synthetases. These data question the concept that the classification of aminoacyl-tRNA synthetases does not vary throughout living systems.

  19. Orthogonal Mapping in Two Dimensions

    NASA Astrophysics Data System (ADS)

    Duraiswami, Ramani; Prosperetti, Andrea

    1992-02-01

    A method for the generation of orthogonal boundary-fitted curvilinear coordinates for arbitrary simply- and doubly-connected domains is developed on the basis of the theory of quasi-conformal mappings of quadrilaterals and of previous work by Ryskin and Leal. The method has useful applications in orthogonal grid generation in two-dimensional and axi-symmetric domains and in the extension of rapid elliptic solvers and spectral methods to complex geometries. A new technique for the calculation of the conformal module of quadrilaterals is also presented.

  20. Orthogonal subspaces for correlation masking

    NASA Technical Reports Server (NTRS)

    Juday, Richard D.; Fisher, Timothy E.

    1990-01-01

    A digital correlation mask that induces orthogonality among a prescribed set of reference imagery is described. In this particular implementation, the resulting correlation is undersampled and shift-variant, though if it is applied to optical correlators, those limitations are removed. A method of introducing orthogonality among the weights among the training set from which filter values are obtained is derived, so that the correlation value from a given filter is representative of the unique nature of the reference object as compared against the other objects in the training class.

  1. Orthogonality catastrophes in quantum electrodynamics

    NASA Astrophysics Data System (ADS)

    Merlin, R.

    2017-02-01

    The insertion of a small polarizable particle in an arbitrarily large optical cavity significantly alters the quantum-mechanical state of the electromagnetic field in that the photon ground state of the empty cavity and that of the cavity with the particle become mutually orthogonal and, thus, cannot be connected adiabatically in the infinite limit. The photon problem can be mapped exactly onto that of a many-body system of fermions, which is known to exhibit an orthogonality catastrophe when a finite-range local potential is introduced. We predict that the motion of polarizable objects inside a cavity as well as their addition and removal from the cavity, will generate a macroscopic, diverging number of low-energy photons. The significance of these results in regard to the quantum measurement problem and the dynamical Casimir effect are also discussed.

  2. Orthogonality preserving infinite dimensional quadratic stochastic operators

    SciTech Connect

    Akın, Hasan; Mukhamedov, Farrukh

    2015-09-18

    In the present paper, we consider a notion of orthogonal preserving nonlinear operators. We introduce π-Volterra quadratic operators finite and infinite dimensional settings. It is proved that any orthogonal preserving quadratic operator on finite dimensional simplex is π-Volterra quadratic operator. In infinite dimensional setting, we describe all π-Volterra operators in terms orthogonal preserving operators.

  3. Peptide Synthetase Gene in Trichoderma virens

    PubMed Central

    Wilhite, S. E.; Lumsden, R. D.; Straney, D. C.

    2001-01-01

    Trichoderma virens (synonym, Gliocladium virens), a deuteromycete fungus, suppresses soilborne plant diseases caused by a number of fungi and is used as a biocontrol agent. Several traits that may contribute to the antagonistic interactions of T. virens with disease-causing fungi involve the production of peptide metabolites (e.g., the antibiotic gliotoxin and siderophores used for iron acquisition). We cloned a 5,056-bp partial cDNA encoding a putative peptide synthetase (Psy1) from T. virens using conserved motifs found within the adenylate domain of peptide synthetases. Sequence similarities with conserved motifs of the adenylation domain, acyl transfer, and two condensation domains support identification of the Psy1 gene as a gene that encodes a peptide synthetase. Disruption of the native Psy1 gene through gene replacement was used to identify the function of this gene. Psy1 disruptants produced normal amounts of gliotoxin but grew poorly under low-iron conditions, suggesting that Psy1 plays a role in siderophore production. Psy1 disruptants cannot produce the major T. virens siderophore dimerum acid, a dipetide of acylated Nδ-hydroxyornithine. Biocontrol activity against damping-off diseases caused by Pythium ultimum and Rhizoctonia solani was not reduced by the Psy1 disruption, suggesting that iron competition through dimerum acid production does not contribute significantly to disease suppression activity under the conditions used. PMID:11679326

  4. Purification of glutathionylspermidine and trypanothione synthetases from Crithidia fasciculata.

    PubMed Central

    Smith, K.; Nadeau, K.; Bradley, M.; Walsh, C.; Fairlamb, A. H.

    1992-01-01

    Two enzymes involved in the biosynthesis of the trypanosomatid-specific dithiol trypanothione-glutathionylspermidine (Gsp) synthetase and trypanothione (TSH) synthetase--have been identified and purified individually from Crithidia fasciculata. The Gsp synthetase has been purified 93-fold and the TSH synthetase 52-fold to apparent homogeneity from a single DEAE fraction that contained both activities. This constitutes the first indication that the enzymatic conversion of two glutathione molecules and one spermidine to the N1,N8-bis(glutathionyl)spermidine (TSH) occurs in two discrete enzymatic steps. Gsp synthetase, which has a kcat of 600/min, shows no detectable TSH synthetase activity, whereas TSH synthetase does not make any detectable Gsp and has a kcat of 75/min. The 90-kDa Gsp synthetase and 82-kDa TSH synthetase are separable on phenyl Superose and remain separated on gel filtration columns in high salt (0.8 M NaCl). Active complexes can be formed under low to moderate salt conditions (0.0-0.15 M NaCl), consistent with a functional complex in vivo. PMID:1304372

  5. Dexamethasone regulates glutamine synthetase expression in rat skeletal muscles

    NASA Technical Reports Server (NTRS)

    Max, Stephen R.; Konagaya, Masaaki; Konagaya, Yoko; Thomas, John W.; Banner, Carl; Vitkovic, Ljubisa

    1986-01-01

    The regulation of glutamine synthetase by glucocorticoids in rat skeletal muscles was studied. Administration of dexamethasone strikingly enhanced glutamine synthetase activity in plantaris and soleus muscles. The dexamethasone-mediated induction of glutamine synthetase activity was blocked to a significant extent by orally administered RU38486, a glucocorticoid antagonist, indicating the involvement of intracellular glucocorticoid receptors in the induction. Northern blot analysis revealed that dexamethasone-mediated enhancement of glutamine synthetase activity involves dramatically increased levels of glutamine synthetase mRNA. The induction of glutamine synthetase was selective in that glutaminase activity of soleus and plantaris muscles was not increased by dexamethasone. Furthermore, dexamethasone treatment resulted in only a small increase in glutamine synthetase activity in the heart. Accordingly, there was only a slight change in glutamine synthetase mRNA level in this tissue. Thus, glucocorticoids regulate glutamine synthetase gene expression in rat muscles at the transcriptional level via interaction with intracellular glutamine production by muscle and to mechanisms underlying glucocorticoid-induced muscle atrophy.

  6. Efficient algorithms for orthogonal packing problems

    NASA Astrophysics Data System (ADS)

    Chekanin, A. V.; Chekanin, V. A.

    2013-10-01

    The NP complete problem of the orthogonal packing of objects of arbitrary dimension is considered in the general form. A new model for representing objects in containers is proposed that ensures the fast design of an orthogonal packing. New heuristics for the placement of orthogonal packing are proposed. A single-pass heuristic algorithm and a multimethod genetic algorithm are developed that optimize an orthogonal packing solution by increasing the packing density. Numerical experiments for two- and three-dimensional orthogonal packing problems are performed.

  7. Supramolecular Polymers with Orthogonal Functionality

    DTIC Science & Technology

    2014-12-11

    and UPy-PEB-UPy assembled in an orthogonal fashion. Additionally, the DMTA showed two transitions for the disassembly of UPy (ca. 60 °C) and Fe2+-Mebip...resulted in a statistical mixture of noncovalent interactions, whereas blends with [Fe(Mebip-PEB- Mebip)](ClO4)2 and UPy-PEB-UPy assembled in an...polymers due to the dynamic nature of noncovalent interactions (π−π, metal−ligand, and hydrogen bonding). The latter serve to assemble the monomeric

  8. Site Directed Mutagenesis of Schizosaccharomyces pombe Glutathione Synthetase Produces an Enzyme with Homoglutathione Synthetase Activity

    PubMed Central

    Dworeck, Tamara; Zimmermann, Martin

    2012-01-01

    Three different His-tagged, mutant forms of the fission yeast glutathione synthetase (GSH2) were derived by site-directed mutagenesis. The mutant and wild-type enzymes were expressed in E. coli DH5α and affinity purified in a two-step procedure. Analysis of enzyme activity showed that it was possible to shift the substrate specificity of GSH2 from Gly (km 0,19; wild-type) to β-Ala or Ser. One mutation (substitution of Ile471, Cy472 to Met and Val and Ala 485 and Thr486 to Leu and Pro) increased the affinity of GSH2 for β-Ala (km 0,07) and lowered the affinity for Gly (km 0,83), which is a characteristic of the enzyme homoglutathione synthetase found in plants. Substitution of Ala485 and Thr486 to Leu and Pro only, increased instead the affinity of GSH2 for Ser (km 0,23) as a substrate, while affinity to Gly was preserved (km 0,12). This provides a new biosynthetic pathway for hydroxymethyl glutathione, which is known to be synthesized from glutathione and Ser in a reaction catalysed by carboxypeptidase Y. The reported findings provide further insight into how specific amino acids positioned in the GSH2 active site facilitate the recognition of different amino acid substrates, furthermore they support the evolutionary theory that homoglutathione synthetase evolved from glutathione synthetase by a single gene duplication event. PMID:23091597

  9. In vivo modification of Azotobacter chroococcum glutamine synthetase.

    PubMed

    Muñoz-Centeno, M C; Cejudo, F J; Paneque, A

    1994-03-15

    A monospecific anti-(glutamine synthetase) antibody raised against glutamine synthetase of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 immunoreacted with glutamine synthetase from the N2-fixing heterotrophic bacterium Azotobacter chroococcum. In Western-blotting experiments this antibody recognized a single protein of a molecular mass of 59 kDa corresponding to glutamine synthetase subunit. This protein was in vivo-labelled in response to addition of ammonium, both [3H]adenine and H(3)32PO4 preincubation of the cells being equally effective. Nevertheless, the amount of glutamine synthetase present in A. chroococcum was independent of the available nitrogen source. Modified, inactive glutamine synthetase was re-activated by treatment with snake-venom phosphodiesterase but not by alkaline phosphatase. L-Methionine-DL-sulphoximine, an inhibitor of glutamine synthetase, prevented the enzyme from being covalently modified. We conclude that, in A. chroococcum, glutamine synthetase is adenylylated in response to ammonium and that for the modification to take place ammonium must be metabolized.

  10. In vivo modification of Azotobacter chroococcum glutamine synthetase.

    PubMed Central

    Muñoz-Centeno, M C; Cejudo, F J; Paneque, A

    1994-01-01

    A monospecific anti-(glutamine synthetase) antibody raised against glutamine synthetase of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 immunoreacted with glutamine synthetase from the N2-fixing heterotrophic bacterium Azotobacter chroococcum. In Western-blotting experiments this antibody recognized a single protein of a molecular mass of 59 kDa corresponding to glutamine synthetase subunit. This protein was in vivo-labelled in response to addition of ammonium, both [3H]adenine and H(3)32PO4 preincubation of the cells being equally effective. Nevertheless, the amount of glutamine synthetase present in A. chroococcum was independent of the available nitrogen source. Modified, inactive glutamine synthetase was re-activated by treatment with snake-venom phosphodiesterase but not by alkaline phosphatase. L-Methionine-DL-sulphoximine, an inhibitor of glutamine synthetase, prevented the enzyme from being covalently modified. We conclude that, in A. chroococcum, glutamine synthetase is adenylylated in response to ammonium and that for the modification to take place ammonium must be metabolized. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7908189

  11. Retinal Vasculitis in Anti-Synthetase Syndrome.

    PubMed

    Donovan, Christopher P; Pecen, Paula E; Baynes, Kimberly; Ehlers, Justis P; Srivastava, Sunil K

    2016-09-01

    A 31-year-old woman with a history of anti-synthetase syndrome-related myositis and interstitial lung disease presented with acute-onset blurry vision and rash on her hands and feet. Visual acuity was hand motion in her right eye and 20/40 in her left eye. Dilated fundus exam showed extensive retinal vasculitis, diffuse intraretinal hemorrhages, and subretinal fluid. Optical coherence tomography revealed significant macular thickening, and fluorescein angiography revealed vascular leakage with peripheral nonperfusion. Aggressive systemic immunosuppression was initiated, with gradual resolution of her disease during 8 months of follow-up. [Ophthalmic Surg Lasers Imaging Retina. 2016;47:874-879.].

  12. Characterization of Cereulide Synthetase, a Toxin-Producing Macromolecular Machine

    PubMed Central

    Alonzo, Diego A.; Magarvey, Nathan A.; Schmeing, T. Martin

    2015-01-01

    Cereulide synthetase is a two-protein nonribosomal peptide synthetase system that produces a potent emetic toxin in virulent strains of Bacillus cereus. The toxin cereulide is a depsipeptide, as it consists of alternating aminoacyl and hydroxyacyl residues. The hydroxyacyl residues are derived from keto acid substrates, which cereulide synthetase selects and stereospecifically reduces with imbedded ketoreductase domains before incorporating them into the growing depsipeptide chain. We present an in vitro biochemical characterization of cereulide synthetase. We investigate the kinetics and side chain specificity of α-keto acid selection, evaluate the requirement of an MbtH-like protein for adenylation domain activity, assay the effectiveness of vinylsulfonamide inhibitors on ester-adding modules, perform NADPH turnover experiments and evaluate in vitro depsipeptide biosynthesis. This work also provides biochemical insight into depsipeptide-synthesizing nonribosomal peptide synthetases responsible for other bioactive molecules such as valinomycin, antimycin and kutzneride. PMID:26042597

  13. Numerical analysis of the orthogonal descent method

    SciTech Connect

    Shokov, V.A.; Shchepakin, M.B.

    1994-11-01

    The author of the orthogonal descent method has been testing it since 1977. The results of these tests have only strengthened the need for further analysis and development of orthogonal descent algorithms for various classes of convex programming problems. Systematic testing of orthogonal descent algorithms and comparison of test results with other nondifferentiable optimization methods was conducted at TsEMI RAN in 1991-1992 using the results.

  14. FRET monitoring of a nonribosomal peptide synthetase.

    PubMed

    Alfermann, Jonas; Sun, Xun; Mayerthaler, Florian; Morrell, Thomas E; Dehling, Eva; Volkmann, Gerrit; Komatsuzaki, Tamiki; Yang, Haw; Mootz, Henning D

    2017-09-01

    Nonribosomal peptide synthetases (NRPSs) are multidomain enzyme templates for the synthesis of bioactive peptides. Large-scale conformational changes during peptide assembly are obvious from crystal structures, yet their dynamics and coupling to catalysis are poorly understood. We have designed an NRPS FRET sensor to monitor, in solution and in real time, the adoption of the productive transfer conformation between phenylalanine-binding adenylation (A) and peptidyl-carrier-protein domains of gramicidin synthetase I from Aneurinibacillus migulanus. The presence of ligands, substrates or intermediates induced a distinct fluorescence resonance energy transfer (FRET) readout, which was pinpointed to the population of specific conformations or, in two cases, mixtures of conformations. A pyrophosphate switch and lysine charge sensors control the domain alternation of the A domain. The phenylalanine-thioester and phenylalanine-AMP products constitute a mechanism of product inhibition and release that is involved in ordered assembly-line peptide biosynthesis. Our results represent insights from solution measurements into the conformational dynamics of the catalytic cycle of NRPSs.

  15. Aminoacyl-tRNA synthetases database.

    PubMed

    Szymanski, M; Deniziak, M A; Barciszewski, J

    2001-01-01

    Aminoacyl-tRNA synthetases (AARSs) are at the center of the question of the origin of life. They constitute a family of enzymes integrating the two levels of cellular organization: nucleic acids and proteins. AARSs arose early in evolution and are believed to be a group of ancient proteins. They are responsible for attaching amino acid residues to their cognate tRNA molecules, which is the first step in the protein synthesis. The role they play in a living cell is essential for the precise deciphering of the genetic code. The analysis of AARSs evolutionary history was not possible for a long time due to a lack of a sufficiently large number of their amino acid sequences. The emerging picture of synthetases' evolution is a result of recent achievements in genomics [Woese,C., Olsen,G.J., Ibba,M. and Söll,D. (2000) Microbiol. Mol. Biol. Rev., 64, 202-236]. In this paper we present a short introduction to the AARSs database. The updated database contains 1047 AARS primary structures from archaebacteria, eubacteria, mitochondria, chloroplasts and eukaryotic cells. It is the compilation of amino acid sequences of all AARSs known to date, which are available as separate entries via the WWW at http://biobases.ibch.poznan.pl/aars/.

  16. The microsomal dicarboxylyl-CoA synthetase.

    PubMed Central

    Vamecq, J; de Hoffmann, E; Van Hoof, F

    1985-01-01

    Dicarboxylic acids are products of the omega-oxidation of monocarboxylic acids. We demonstrate that in rat liver dicarboxylic acids (C5-C16) can be converted into their CoA esters by a dicarboxylyl-CoA synthetase. During this activation ATP, which cannot be replaced by GTP, is converted into AMP and PPi, both acting as feedback inhibitors of the reaction. Thermolabile at 37 degrees C, and optimally active at pH 6.5, dicarboxylyl-CoA synthetase displays the highest activity on dodecanedioic acid (2 micromol/min per g of liver). Cell-fractionation studies indicate that this enzyme belongs to the hepatic microsomal fraction. Investigations about the fate of dicarboxylyl-CoA esters disclosed the existence of an oxidase, which could be measured by monitoring the production of H2O2. In our assay conditions this H2O2 production is dependent on and closely follows the CoA consumption. It appears that the chain-length specificity of the handling of dicarboxylic acids by this catabolic pathway (activation to acyl-CoA and oxidation with H2O2 production) parallels the pattern of the degradation of exogenous dicarboxylic acids in vivo. PMID:4062873

  17. Orthogonal coding of object location.

    PubMed

    Knutsen, Per Magne; Ahissar, Ehud

    2009-02-01

    It has been argued whether internal representations are encoded using a universal ('the neural code') or multiple codes. Here, we review a series of experiments that demonstrate that tactile encoding of object location via whisking employs an orthogonal, triple-code scheme. Rats, and other rodents, actively move the whiskers back and forth to localize and identify objects. Neural recordings from primary sensory afferents, along with behavioral observations, demonstrate that vertical coordinates of contacted objects are encoded by the identity of activated afferents, horizontal coordinates by the timing of activation and radial coordinates by the intensity of activation. Because these codes are mutually independent, the three-dimensional location of an object could, in principle, be encoded by individual afferents during single whisker-object contacts. One advantage of such a same-neuron-different-codes scheme over the traditionally assumed same-code-different-neurons scheme is a reduction of code ambiguity that, in turn, simplifies decoding circuits.

  18. Orthogonal separations: Comparison of orthogonality metrics by statistical analysis.

    PubMed

    Schure, Mark R; Davis, Joe M

    2015-10-02

    Twenty orthogonality metrics (OMs) derived from convex hull, information theory, fractal dimension, correlation coefficients, nearest neighbor distances and bin-density techniques were calculated from a diverse group of 47 experimental two-dimensional (2D) chromatograms. These chromatograms comprise two datasets; one dataset is a collection of 2D chromatograms from Peter Carr's laboratory at the University of Minnesota, and the other dataset is based on pairs of one-dimensional chromatograms previously published by Martin Gilar and coworkers (Waters Corp.). The chromatograms were pooled to make a third or combined dataset. Cross-correlation results suggest that specific OMs are correlated within families of nearest neighbor methods, correlation coefficients and the information theory methods. Principal component analysis of the OMs show that none of the OMs stands out as clearly better at explaining the data variance than any another OM. Principal component analysis of individual chromatograms shows that different OMs favor certain chromatograms. The chromatograms exhibit a range of quality, as subjectively graded by nine experts experienced in 2D chromatography. The subjective (grading) evaluations were taken at two intervals per expert and demonstrated excellent consistency for each expert. Excellent agreement for both very good and very bad chromatograms was seen across the range of experts. However, evaluation uncertainty increased for chromatograms that were judged as average to mediocre. The grades were converted to numbers (percentages) for numerical computations. The percentages were correlated with OMs to establish good OMs for evaluating the quality of 2D chromatograms. Certain metrics correlate better than others. However, these results are not consistent across all chromatograms examined. Most of the nearest neighbor methods were observed to correlate poorly with the percentages. However, one method, devised by Clark and Evans, appeared to work

  19. Non-orthogonal subband/transform coder

    NASA Technical Reports Server (NTRS)

    Glover, Daniel R. (Inventor)

    1993-01-01

    The present invention is directed to a simplified digital subband coder/decoder. In the present invention a signal is fed into a coder. The coder uses a non-orthogonal algorithm that is simply implemented in the coder hardware. The simple non-orthogonal design is then used in the implementation of the decoder to decode the signal.

  20. Glutamine Synthetase: Role in Neurological Disorders.

    PubMed

    Jayakumar, Arumugam R; Norenberg, Michael D

    2016-01-01

    Glutamine synthetase (GS) is an ATP-dependent enzyme found in most species that synthesizes glutamine from glutamate and ammonia. In brain, GS is exclusively located in astrocytes where it serves to maintain the glutamate-glutamine cycle, as well as nitrogen metabolism. Changes in the activity of GS, as well as its gene expression, along with excitotoxicity, have been identified in a number of neurological conditions. The literature describing alterations in the activation and gene expression of GS, as well as its involvement in different neurological disorders, however, is incomplete. This review summarizes changes in GS gene expression/activity and its potential contribution to the pathogenesis of several neurological disorders, including hepatic encephalopathy, ischemia, epilepsy, Alzheimer's disease, amyotrophic lateral sclerosis, traumatic brain injury, Parkinson's disease, and astroglial neoplasms. This review also explores the possibility of targeting GS in the therapy of these conditions.

  1. Mechanistic issues in asparagine synthetase catalysis.

    PubMed

    Richards, N G; Schuster, S M

    1998-01-01

    The enzymatic synthesis of asparagine is an ATP-dependent process that utilizes the nitrogen atom derived from either glutamine or ammonia. Despite a long history of kinetic and mechanistic investigation, there is no universally accepted catalytic mechanism for this seemingly straightforward carboxyl group activating enzyme, especially as regards those steps immediately preceding amide bond formation. This chapter considers four issues dealing with the mechanism: (a) the structural organization of the active site(s) partaking in glutamine utilization and aspartate activation; (b) the relationship of asparagine synthetase to other amidotransferases; (c) the way in which ATP is used to activate the beta-carboxyl group; and (d) the detailed mechanism by which nitrogen is transferred.

  2. Cosmology with orthogonal nilpotent superfields

    NASA Astrophysics Data System (ADS)

    Ferrara, Sergio; Kallosh, Renata; Thaler, Jesse

    2016-02-01

    We study the application of a supersymmetric model with two constrained supermultiplets to inflationary cosmology. The first superfield S is a stabilizer chiral superfield satisfying a nilpotency condition of degree 2, S2=0 . The second superfield Φ is the inflaton chiral superfield, which can be combined into a real superfield B ≡1/2 i (Φ -Φ ¯ ) . The real superfield B is orthogonal to S , S B =0 , and satisfies a nilpotency condition of degree 3, B3=0 . We show that these constraints remove from the spectrum the complex scalar sgoldstino, the real scalar inflaton partner (i.e. the "sinflaton"), and the fermionic inflatino. The corresponding supergravity model with de Sitter vacua describes a graviton, a massive gravitino, and one real scalar inflaton, with both the goldstino and inflatino being absent in unitary gauge. We also discuss relaxed superfield constraints where S2=0 and S Φ ¯ is chiral, which removes the sgoldstino and inflatino, but leaves the sinflaton in the spectrum. The cosmological model building in both of these inflatino-less models offers some advantages over existing constructions.

  3. Which are more accurate, orthogonal or non-orthogonal sonic anemometers?

    NASA Astrophysics Data System (ADS)

    Massman, W. J.; Frank, J. M.; Swiatek, E.; Zimmerman, H.; Ewers, B. E.

    2013-12-01

    Sonic anemometry is fundamental to all eddy-covariance studies of surface energy, ecosystem carbon, and water balance. Recent studies have shown the potential underestimation of the vertical wind fluctuations among the most commonly encountered anemometer models, but thus far testing has been focused on non-orthogonal sonic anemometer designs. We hypothesize that these underestimates are systematic to the non-orthogonal design and not attributable to a single manufacturer. If so, orthogonal measurements of vertical wind should be more accurate. We tested this by conducting an experiment to measure the relative consistency between vertical and horizontal wind measurements for three sonic anemometer designs: orthogonal, non-orthogonal, and quasi-orthogonal. Both the orthogonal and non-orthogonal models were from a single manufacturer (K-probe and A-probe, Applied Technologies, Inc.) while the quasi-orthogonal design featured non-orthogonal u- and v-axes but with an orthogonal w-axis (CSAT3V, Campbell Scientific, Inc.). We conducted a 12-week experiment, testing four sonic anemometers relative to a control (CSAT3, Campbell Scientific, Inc.), each week randomly selecting at least one of each model from a pool of twelve instruments (three of each model) and randomly locating the test anemometers around the control. Half-way through the week the test anemometers were re-mounted in a horizontal position. Work was done at the GLEES AmeriFlux site (southeastern Wyoming, USA) which experiences large, uni-directional wind and turbulence. Results are discussed.

  4. TRYPTOPHANASE-TRYPTOPHAN SYNTHETASE SYSTEMS IN ESCHERICHIA COLI III.

    PubMed Central

    Freundlich, Martin; Lichstein, Herman C.

    1962-01-01

    Freundlich, Martin (University of Minnesota, Minneapolis) and Herman C. Lichstein. Tryptophanase-tryptophan synthetase systems in Escherichia coli. III. Requirements for enzyme synthesis. J. Bacteriol. 84:996–1006. 1962.—The requirements for the formation of tryptophanase and tryptophan synthetase in Escherichia coli during repression release were studied. The kinetics of the formation of tryptophan synthetase differed in the two strains examined; this was attributed to differences in the endogenous level of tryptophan in the bacterial cells. The formation of both enzymes was inhibited by chloramphenicol, and by the absence of arginine in an arginine-requiring mutant. These results are indicative of a requirement for protein synthesis for enzyme formation. Requirements for nucleic acid synthesis were examined by use of a uracil- and thymine-requiring mutant, and with purine and pyrimidine analogues. The results obtained suggest that some type of ribonucleic acid synthesis was necessary for the formation of tryptophanase and tryptophan synthetase. PMID:13959620

  5. [Orthogonal Vector Projection Algorithm for Spectral Unmixing].

    PubMed

    Song, Mei-ping; Xu, Xing-wei; Chang, Chein-I; An, Ju-bai; Yao, Li

    2015-12-01

    Spectrum unmixing is an important part of hyperspectral technologies, which is essential for material quantity analysis in hyperspectral imagery. Most linear unmixing algorithms require computations of matrix multiplication and matrix inversion or matrix determination. These are difficult for programming, especially hard for realization on hardware. At the same time, the computation costs of the algorithms increase significantly as the number of endmembers grows. Here, based on the traditional algorithm Orthogonal Subspace Projection, a new method called. Orthogonal Vector Projection is prompted using orthogonal principle. It simplifies this process by avoiding matrix multiplication and inversion. It firstly computes the final orthogonal vector via Gram-Schmidt process for each endmember spectrum. And then, these orthogonal vectors are used as projection vector for the pixel signature. The unconstrained abundance can be obtained directly by projecting the signature to the projection vectors, and computing the ratio of projected vector length and orthogonal vector length. Compared to the Orthogonal Subspace Projection and Least Squares Error algorithms, this method does not need matrix inversion, which is much computation costing and hard to implement on hardware. It just completes the orthogonalization process by repeated vector operations, easy for application on both parallel computation and hardware. The reasonability of the algorithm is proved by its relationship with Orthogonal Sub-space Projection and Least Squares Error algorithms. And its computational complexity is also compared with the other two algorithms', which is the lowest one. At last, the experimental results on synthetic image and real image are also provided, giving another evidence for effectiveness of the method.

  6. On orthogonality preserving quadratic stochastic operators

    SciTech Connect

    Mukhamedov, Farrukh; Taha, Muhammad Hafizuddin Mohd

    2015-05-15

    A quadratic stochastic operator (in short QSO) is usually used to present the time evolution of differing species in biology. Some quadratic stochastic operators have been studied by Lotka and Volterra. In the present paper, we first give a simple characterization of Volterra QSO in terms of absolutely continuity of discrete measures. Further, we introduce a notion of orthogonal preserving QSO, and describe such kind of operators defined on two dimensional simplex. It turns out that orthogonal preserving QSOs are permutations of Volterra QSO. The associativity of genetic algebras generated by orthogonal preserving QSO is studied too.

  7. An orthogonalized platform for genetic code expansion in both bacteria and eukaryotes.

    PubMed

    Italia, James S; Addy, Partha Sarathi; Wrobel, Chester J J; Crawford, Lisa A; Lajoie, Marc J; Zheng, Yunan; Chatterjee, Abhishek

    2017-02-13

    In this study, we demonstrate the feasibility of expanding the genetic code of Escherichia coli using its own tryptophanyl-tRNA synthetase and tRNA (TrpRS-tRNA(Trp)) pair. This was made possible by first functionally replacing this endogenous pair with an E. coli-optimized counterpart from Saccharomyces cerevisiae, and then reintroducing the liberated E. coli TrpRS-tRNA(Trp) pair into the resulting strain as a nonsense suppressor, which was then followed by its directed evolution to genetically encode several new unnatural amino acids (UAAs). These engineered TrpRS-tRNA(Trp) variants were also able to drive efficient UAA mutagenesis in mammalian cells. Since bacteria-derived aminoacyl-tRNA synthetase (aaRS)-tRNA pairs are typically orthogonal in eukaryotes, our work provides a general strategy to develop additional aaRS-tRNA pairs that can be used for UAA mutagenesis of proteins expressed in both E. coli and eukaryotes.

  8. Purification and properties of glutamine synthetase from spinach leaves.

    PubMed

    Ericson, M C

    1985-12-01

    The chloroplastic glutamine synthetase of spinach leaves has been purified to homogeneity using affinity chromatography. This involves a tandem ;reactive blue A-agarose' and ;reactive red-A-agarose' as the final step in the procedure. This procedure results in a yield of 18 milligrams of pure glutamine synthetase per kilogram of starting material. The purity of our enzyme has been demonstrated on both one- and two-dimensional polyacrylamide gels.Purified glutamine synthetase has a molecular weight of 360,000 daltons and consists of eight 44,000 dalton subunits. The K(m) is 6.7 millimolar for glutamate, 1.8 millimolar for ATP (synthetase assay), and 37.6 millimolar for glutamine (transferase assay). The isoelectric point is 6.5 and the pH optima are 7.3 in the synthetase assay and 6.4 in the transferase assay. The irreversible, competitive inhibitors methionine sulfoxamine and phosphinothricin have K(i) values of 0.1 millimolar and 6.1 micromolar, respectively. Amino acid analysis has been carried out and the results compared with published analyses for other isoforms of glutamine synthetase.

  9. TRYPTOPHANASE-TRYPTOPHAN SYNTHETASE SYSTEMS IN ESCHERICHIA COLI I.

    PubMed Central

    Freundlich, Martin; Lichstein, Herman C.

    1962-01-01

    Freundlich, Martin (University of Minnesota, Minneapolis) and Herman C. Lichstein. Tryptophanase-tryptophan synthetase systems in Escherichia coli. I. Effect of tryptophan and related compounds. J. Bacteriol. 84:979–987. 1962.—The effect of tryptophan and related compounds on tryptophanase and tryptophan synthetase formation in Escherichia coli was determined. Several of these compounds stimulated the formation of tryptophanase while concomitantly decreasing the production of synthetase. A number of tryptophan analogues were found to inhibit growth. The possible mode of action of these substances was examined further. 5-Hydroxytryptophan greatly inhibited the formation of synthetase and also reduced growth. Its inhibitory action on growth was attributed, at least partially, to the false feedback inhibition of anthranilic acid formation. Tryptamine was found to be a potent inhibitor of the activity of synthetase, as well as of the enzyme(s) involved in the synthesis of anthranilic acid from shikimic acid. However, growth reduction was only partially reversed by tryptophan. Indole-3-acetic acid and indole-3-propionic acid decreased growth and increased the formation of synthetase six- to eightfold. The action of these compounds was ascribed to their ability to block the endogenous formation of tryptophan. PMID:13959621

  10. Evolution of aminoacyl-tRNA synthetase quaternary structure and activity: Saccharomyces cerevisiae mitochondrial phenylalanyl-tRNA synthetase.

    PubMed Central

    Sanni, A; Walter, P; Boulanger, Y; Ebel, J P; Fasiolo, F

    1991-01-01

    Phenylalanyl-tRNA synthetases [L-phenylalanine:tRNAPhe ligase (AMP-forming), EC 6.1.1.20] from Escherichia coli, yeast cytoplasm, and mammalian cytoplasm have an unusual conserved alpha 2 beta 2 quaternary structure that is shared by only one other aminoacyl-tRNA synthetase. Both subunits are required for activity. We show here that a single mitochondrial polypeptide from Saccharomyces cerevisiae is an active phenylalanyl-tRNA synthetase. This protein (the MSF1 gene product) is active as a monomer. It has all three characteristic sequence motifs of the class II aminoacyl-tRNA synthetases, and its activity may result from the recruitment of additional sequences into an alpha-subunit-like structure. Images PMID:1924298

  11. Gaussian quadrature for multiple orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Coussement, Jonathan; van Assche, Walter

    2005-06-01

    We study multiple orthogonal polynomials of type I and type II, which have orthogonality conditions with respect to r measures. These polynomials are connected by their recurrence relation of order r+1. First we show a relation with the eigenvalue problem of a banded lower Hessenberg matrix Ln, containing the recurrence coefficients. As a consequence, we easily find that the multiple orthogonal polynomials of type I and type II satisfy a generalized Christoffel-Darboux identity. Furthermore, we explain the notion of multiple Gaussian quadrature (for proper multi-indices), which is an extension of the theory of Gaussian quadrature for orthogonal polynomials and was introduced by Borges. In particular, we show that the quadrature points and quadrature weights can be expressed in terms of the eigenvalue problem of Ln.

  12. Orthogonal Chirp-Based Ultrasonic Positioning

    PubMed Central

    Khyam, Mohammad Omar; Ge, Shuzhi Sam; Li, Xinde; Pickering, Mark

    2017-01-01

    This paper presents a chirp based ultrasonic positioning system (UPS) using orthogonal chirp waveforms. In the proposed method, multiple transmitters can simultaneously transmit chirp signals, as a result, it can efficiently utilize the entire available frequency spectrum. The fundamental idea behind the proposed multiple access scheme is to utilize the oversampling methodology of orthogonal frequency-division multiplexing (OFDM) modulation and orthogonality of the discrete frequency components of a chirp waveform. In addition, the proposed orthogonal chirp waveforms also have all the advantages of a classical chirp waveform. Firstly, the performance of the waveforms is investigated through correlation analysis and then, in an indoor environment, evaluated through simulations and experiments for ultrasonic (US) positioning. For an operational range of approximately 1000 mm, the positioning root-mean-square-errors (RMSEs) &90% error were 4.54 mm and 6.68 mm respectively. PMID:28448454

  13. Glucocorticoid receptor-mediated induction of glutamine synthetase in skeletal muscle cells in vitro

    NASA Technical Reports Server (NTRS)

    Max, Stephen R.; Thomas, John W.; Banner, Carl; Vitkovic, Ljubisa; Konagaya, Masaaki

    1987-01-01

    The regulation by glucocorticoids of glutamine synthetase in L6 muscle cells in culture is studied. Glutamine synthetase activity was strikingly enhanced by dexamethasone. The dexamethasone-mediated induction of glutamine synthetase activity was blocked by RU38486, a glucocorticoid antagonist, indicating the involvement of intracellular glucocorticoid receptors in the induction process. RU38486 alone was without effect. Northern blot analysis revealed that dexamethasone-mediated enhancement of glutamine synthetase activity involves increased levels of glutamine synthetase mRNA. Glucocorticoids regulate the expression of glutamine synthetase mRNA in cultured muscle cells via interaction with intracellular receptors. Such regulation may be relevant to control of glutamine production by muscle.

  14. Glucocorticoid receptor-mediated induction of glutamine synthetase in skeletal muscle cells in vitro

    NASA Technical Reports Server (NTRS)

    Max, Stephen R.; Thomas, John W.; Banner, Carl; Vitkovic, Ljubisa; Konagaya, Masaaki

    1987-01-01

    The regulation by glucocorticoids of glutamine synthetase in L6 muscle cells in culture is studied. Glutamine synthetase activity was strikingly enhanced by dexamethasone. The dexamethasone-mediated induction of glutamine synthetase activity was blocked by RU38486, a glucocorticoid antagonist, indicating the involvement of intracellular glucocorticoid receptors in the induction process. RU38486 alone was without effect. Northern blot analysis revealed that dexamethasone-mediated enhancement of glutamine synthetase activity involves increased levels of glutamine synthetase mRNA. Glucocorticoids regulate the expression of glutamine synthetase mRNA in cultured muscle cells via interaction with intracellular receptors. Such regulation may be relevant to control of glutamine production by muscle.

  15. Glutamine synthetase induced spinal seizures in rats.

    PubMed

    Shin, Dong Won; Yoon, Young Sul; Matsumoto, Masato; Huang, Wencheng; Ceraulo, Phil; Young, Wise

    2003-02-01

    Glutamine synthetase (GS) is a key enzyme in the regulation of glutamate neurotransmission in the central nervous system. It is responsible for converting glutamate to glutamine, consuming one ATP and NH3 in the process. Glutamate is neurotoxic when it accumulates in extracellular fluids. We investigated the effects of GS in both a spinal cord injury (SCI) model and normal rats. 0.1-ml of low (2- micro M) and high (55- micro M) concentrations of GS were applied, intrathecally, to the spinal cord of rats under pentobarbital anesthesia. Immediately after an intrathecal injection into the L1-L3 space, the rats developed convulsive movements. These movements initially consisted of myoclonic twitches of the paravertebral muscles close to the injection site, repeated tonic and clonic contractions and extensions of the hind limbs (hind limb seizures) that spread to the fore limbs, and finally rotational axial movements of the body. An EMG of the paravertebral muscles, fore and hind limbs, showed the extent of the muscle activities. GS (2- micro M) caused spinal seizures in the rats after the SCI, and GS (6- micro M) produced seizures in the uninjured anesthetized rats. Denatured GS (70 degrees C, 1 hour) also produced spinal seizures, although higher concentrations were required. We suggest that GS may be directly blocking the release of GABA, or the receptors, in the spinal cord.

  16. Chemical modification of E. coli glutamine synthetase

    SciTech Connect

    DiIanni, C.L.; Colanduoni, J.A.; Collins, R.; Villafranca, J.J.

    1986-05-01

    Thiourea trioxide partially inactivates E. coli glutamine synthetase (GS) (approx.25%) by reacting only with lysine residues, producing homoarginine. Thiourea dioxide totally inactivates GS by reacting with both lysine and histidine residues. The K/sub m/ values for thiourea trioxide modified enzyme are 0.21 mM for ATP and 10 mM for glutamate which are about threefold higher than for native GS. Using (/sup 14/C) thiourea trioxide, 2.3 +/- 0.2 moles of reagent were incorporated per monomer. The same number of homoarginine residues were found by amino acid analysis. Modification of GS with hydroxylamine results in total inactivation resulting from reaction with histidine. Fluorescence titrations indicate that substrate binding to the modified enzyme is weaker than to the native enzyme. EPR spectra of bound Mn/sup 2 +/ indicate that metal ion binding is unaffected by hydroxylamine modification. However, metal ion binding is weaker to the modified enzyme. Protection from hydroxylamine inactivation is observed with ATP + Glutamate, AMPPNP + Glutamate, and MgCl/sub 2/.

  17. Dihydrofolate synthetase and folylpolyglutamate synthetase: direct evidence for intervention of acyl phosphate intermediates

    SciTech Connect

    Banerjee, R.V.; Shane, B.; McGuire, J.J.; Coward, J.K.

    1988-12-13

    The transfer of /sup 17/O and/or /sup 18/O from (COOH-/sup 17/O or -/sup 18/O) enriched substrates to inorganic phosphate (P/sub i/) has been demonstrated for two enzyme-catalyzed reactions involved in folate biosynthesis and glutamylation. COOH-/sup 18/O-labeled folate, methotrexate, and dihydropteroate, in addition to (/sup 17/O)-glutamate, were synthesized and used as substrates for folylpolyglutamate synthetase (FPGS) isolated from Escherichia coli, hog liver, and rat liver and for dihydrofolate synthetase (DHFS) isolated from E. coli. P/sub i/ was purified from the reaction mixtures and converted to trimethyl phosphate (TMP), which was then analyzed for /sup 17/O and /sup 18/O enrichment by nuclear magnetic resonance (NMR) spectroscopy and/or mass spectroscopy. In the reactions catalyzed by the E. coli enzymes, both NMR and quantitative mass spectral analyses established that transfer of the oxygen isotope from the substrate /sup 18/O-enriched carboxyl group to P/sub i/ occurred, thereby providing strong evidence for an acyl phosphate intermediate in both the FPGS- and DHFS-catalyzed reactions. Similar oxygen-transfer experiments were carried out by use of two mammalian enzymes. The small amounts of P/sub i/ obtained from reactions catalyzed by these less abundant FPGS proteins precluded the use of NMR techniques. However, mass spectral analysis of the TMP derived from the mammalian FPGS-catalyzed reactions showed clearly that /sup 18/O transfer had occurred.

  18. Aminoacyl tRNA synthetases and their connections to disease.

    PubMed

    Park, Sang Gyu; Schimmel, Paul; Kim, Sunghoon

    2008-08-12

    Aminoacylation of transfer RNAs establishes the rules of the genetic code. The reactions are catalyzed by an ancient group of 20 enzymes (one for each amino acid) known as aminoacyl tRNA synthetases (AARSs). Surprisingly, the etiology of specific diseases-including cancer, neuronal pathologies, autoimmune disorders, and disrupted metabolic conditions-is connected to specific aminoacyl tRNA synthetases. These connections include heritable mutations in the genes for tRNA synthetases that are causally linked to disease, with both dominant and recessive disease-causing mutations being annotated. Because some disease-causing mutations do not affect aminoacylation activity or apparent enzyme stability, the mutations are believed to affect functions that are distinct from aminoacylation. Examples include enzymes that are secreted as procytokines that, after activation, operate in pathways connected to the immune system or angiogenesis. In addition, within cells, synthetases form multiprotein complexes with each other or with other regulatory factors and in that way control diverse signaling pathways. Although much has been uncovered in recent years, many novel functions, disease connections, and interpathway connections of tRNA synthetases have yet to be worked out.

  19. Functional linkage between the glutaminase and synthetase domains of carbamoyl-phosphate synthetase. Role of serine 44 in carbamoyl-phosphate synthetase-aspartate carbamoyltransferase-dihydroorotase (cad).

    PubMed

    Hewagama, A; Guy, H I; Vickrey, J F; Evans, D R

    1999-10-01

    Mammalian carbamoyl-phosphate synthetase is part of carbamoyl-phosphate synthetase-aspartate carbamoyltransferase-dihydroorotase (CAD), a multifunctional protein that also catalyzes the second and third steps of pyrimidine biosynthesis. Carbamoyl phosphate synthesis requires the concerted action of the glutaminase (GLN) and carbamoyl-phosphate synthetase domains of CAD. There is a functional linkage between these domains such that glutamine hydrolysis on the GLN domain does not occur at a significant rate unless ATP and HCO(3)(-), the other substrates needed for carbamoyl phosphate synthesis, bind to the synthetase domain. The GLN domain consists of catalytic and attenuation subdomains. In the separately cloned GLN domain, the catalytic subdomain is down-regulated by interactions with the attenuation domain, a process thought to be part of the functional linkage. Replacement of Ser(44) in the GLN attenuation domain with alanine increases the k(cat)/K(m) for glutamine hydrolysis 680-fold. The formation of a functional hybrid between the mammalian Ser(44) GLN domain and the Escherichia coli carbamoyl-phosphate synthetase large subunit had little effect on glutamine hydrolysis. In contrast, ATP and HCO(3)(-) did not stimulate the glutaminase activity, indicating that the interdomain linkage had been disrupted. In accord with this interpretation, the rate of glutamine hydrolysis and carbamoyl phosphate synthesis were no longer coordinated. Approximately 3 times more glutamine was hydrolyzed by the Ser(44) --> Ala mutant than that needed for carbamoyl phosphate synthesis. Ser(44), the only attenuation subdomain residue that extends into the GLN active site, appears to be an integral component of the regulatory circuit that phases glutamine hydrolysis and carbamoyl phosphate synthesis.

  20. Haunted Quantum Entanglement, Quantum Erasure, and Orthogonality

    NASA Astrophysics Data System (ADS)

    Snyder, Douglas

    2010-02-01

    Both haunted quantum entanglement (hqe) and quantum erasure (qe) demonstrate interference. For interference, overlapping waves are needed which are likely supplied by equations such as 1/√ 2 [\\vert P/u>+\\vert P/l>] = \\vert P/s> and 1/√ 2 [\\vert P/u>-\\vert P/l>] = \\vert P/a> where \\vert P/u> and \\vert P/l> are generally considered orthogonal (i.e., no overlap) and \\vert P/s> and \\vert P/a> are symmetric and anti-symmetric wave functions. The conventional consideration of orthogonality in hqe and qe may need adjustment given empirical support for the presence of fringes and anti-fringes in qe. Orthogonality as regards hqe and qe is tied to the possibility of obtaining which way information. If this possibility is lost, it would appear that orthogonality which is based on this possibility may be lost. A completed measurement appears central to establishing orthogonality as regards hqe and qe. In hqe, this completed measurement could be for example an atom passing through a double slit arrangement after having emitted a photon in one of two micromaser cavities, thus providing general which way information without specifying through which specific slit the atom passed. In qe, the completed measurement could be this atom subsequently striking a detection screen, providing the ability to obtain information regarding through which specific slit the atom passed. Hqe and qe occur when which way information is lost before their respective completed measurements are made. )

  1. Face hallucination using orthogonal canonical correlation analysis

    NASA Astrophysics Data System (ADS)

    Zhou, Huiling; Lam, Kin-Man

    2016-05-01

    A two-step face-hallucination framework is proposed to reconstruct a high-resolution (HR) version of a face from an input low-resolution (LR) face, based on learning from LR-HR example face pairs using orthogonal canonical correlation analysis (orthogonal CCA) and linear mapping. In the proposed algorithm, face images are first represented using principal component analysis (PCA). Canonical correlation analysis (CCA) with the orthogonality property is then employed, to maximize the correlation between the PCA coefficients of the LR and the HR face pairs to improve the hallucination performance. The original CCA does not own the orthogonality property, which is crucial for information reconstruction. We propose using orthogonal CCA, which is proven by experiments to achieve a better performance in terms of global face reconstruction. In addition, in the residual-compensation process, a linear-mapping method is proposed to include both the inter- and intrainformation about manifolds of different resolutions. Compared with other state-of-the-art approaches, the proposed framework can achieve a comparable, or even better, performance in terms of global face reconstruction and the visual quality of face hallucination. Experiments on images with various parameter settings and blurring distortions show that the proposed approach is robust and has great potential for real-world applications.

  2. The glutamine synthetase gene family in Populus

    PubMed Central

    2011-01-01

    Background Glutamine synthetase (GS; EC: 6.3.1.2, L-glutamate: ammonia ligase ADP-forming) is a key enzyme in ammonium assimilation and metabolism of higher plants. The current work was undertaken to develop a more comprehensive understanding of molecular and biochemical features of GS gene family in poplar, and to characterize the developmental regulation of GS expression in various tissues and at various times during the poplar perennial growth. Results The GS gene family consists of 8 different genes exhibiting all structural and regulatory elements consistent with their roles as functional genes. Our results indicate that the family members are organized in 4 groups of duplicated genes, 3 of which code for cytosolic GS isoforms (GS1) and 1 which codes for the choroplastic GS isoform (GS2). Our analysis shows that Populus trichocarpa is the first plant species in which it was observed the complete GS family duplicated. Detailed expression analyses have revealed specific spatial and seasonal patterns of GS expression in poplar. These data provide insights into the metabolic function of GS isoforms in poplar and pave the way for future functional studies. Conclusions Our data suggest that GS duplicates could have been retained in order to increase the amount of enzyme in a particular cell type. This possibility could contribute to the homeostasis of nitrogen metabolism in functions associated to changes in glutamine-derived metabolic products. The presence of duplicated GS genes in poplar could also contribute to diversification of the enzymatic properties for a particular GS isoform through the assembly of GS polypeptides into homo oligomeric and/or hetero oligomeric holoenzymes in specific cell types. PMID:21867507

  3. Changes in the activity levels of glutamine synthetase, glutaminase and glycogen synthetase in rats subjected to hypoxic stress

    NASA Astrophysics Data System (ADS)

    Vats, P.; Mukherjee, A. K.; Kumria, M. M. L.; Singh, S. N.; Patil, S. K. B.; Rangnathan, S.; Sridharan, K.

    Exposure to high altitude causes loss of body mass and alterations in metabolic processes, especially carbohydrate and protein metabolism. The present study was conducted to elucidate the role of glutamine synthetase, glutaminase and glycogen synthetase under conditions of chronic intermittent hypoxia. Four groups, each consisting of 12 male albino rats (Wistar strain), were exposed to a simulated altitude of 7620 m in a hypobaric chamber for 6 h per day for 1, 7, 14 and 21 days, respectively. Blood haemoglobin, blood glucose, protein levels in the liver, muscle and plasma, glycogen content, and glutaminase, glutamine synthetase and glycogen synthetase activities in liver and muscle were determined in all groups of exposed and in a group of unexposed animals. Food intake and changes in body mass were also monitored. There was a significant reduction in body mass (28-30%) in hypoxia-exposed groups as compared to controls, with a corresponding decrease in food intake. There was rise in blood haemoglobin and plasma protein in response to acclimatisation. Over a three-fold increase in liver glycogen content was observed following 1 day of hypoxic exposure (4.76+/-0.78 mg.g-1 wet tissue in normal unexposed rats; 15.82+/-2.30 mg.g-1 wet tissue in rats exposed to hypoxia for 1 day). This returned to normal in later stages of exposure. However, there was no change in glycogen synthetase activity except for a decrease in the 21-days hypoxia-exposed group. There was a slight increase in muscle glycogen content in the 1-day exposed group which declined significantly by 56.5, 50.6 and 42% following 7, 14, and 21 days of exposure, respectively. Muscle glycogen synthetase activity was also decreased following 21 days of exposure. There was an increase in glutaminase activity in the liver and muscle in the 7-, 14- and 21-day exposed groups. Glutamine synthetase activity was higher in the liver in 7- and 14-day exposed groups; this returned to normal following 21 days of exposure

  4. tRNA synthetase: tRNA Aminoacylation and beyond

    PubMed Central

    Pang, Yan Ling Joy; Poruri, Kiranmai; Martinis, Susan A.

    2014-01-01

    The aminoacyl-tRNA synthetases are prominently known for their classic function in the first step of protein synthesis, where they bear the responsibility of setting the genetic code. Each enzyme is exquisitely adapted to covalently link a single standard amino acid to its cognate set of tRNA isoacceptors. These ancient enzymes have evolved idiosyncratically to host alternate activities that go far beyond their aminoacylation role and impact a wide range of other metabolic pathways and cell signaling processes. The family of aminoacyl-tRNA synthetases have also been suggested as a remarkable scaffold to incorporate new domains that would drive evolution and the emergence of new organisms with more complex function. Because they are essential, the tRNA synthetases have served as pharmaceutical targets for drug and antibiotic development. The recent unfolding of novel important functions for this family of proteins offers new and promising pathways for therapeutic development to treat diverse human diseases. PMID:24706556

  5. Heterogeneity of Glutamine Synthetase Polypeptides in Phaseolus vulgaris L. 1

    PubMed Central

    Lara, Miguel; Porta, Helena; Padilla, Jaime; Folch, Jorge; Sánchez, Federico

    1984-01-01

    Glutamine synthetases from roots, nodules, and leaves of Phaseolus vulgaris L. have been purified to homogeneity and their polypeptide composition determined. The leaf enzyme is composed of six polypeptides. The cytosolic fraction contains two 43,000 dalton polypeptides and the chloroplastic enzyme is formed by four 45,000 dalton polypeptides. Root glutamine synthetase consists only of the same two polypeptides of 43,000 dalton that are present in the leaf enzyme. The nodule enzyme is formed by two polypeptides of 43,000 dalton, one is common to the leaf and root enzyme but the other is specific for N2-fixing nodule tissue. The two glutamine synthetase forms of the nodule contain a different proportion of the 43,000 dalton polypeptides. Images Fig. 1 Fig. 2 Fig. 4 PMID:16663942

  6. Inhibition of Plant Glutamine Synthetases by Substituted Phosphinothricins

    PubMed Central

    Logusch, Eugene W.; Walker, Daniel M.; McDonald, John F.; Franz, John E.

    1991-01-01

    Glutamine synthetase (GS) utilizes various substituted glutamic acids as substrates. We have used this information to design herbicidal α- and γ-substituted analogs of phosphinothricin (l-2-amino-4-(hydroxymethylphosphinyl)butanoic acid, PPT), a naturally occurring GS inhibitor and a potent herbicide. The substituted phosphinothricins inhibit cytosolic sorghum GS1 and chloroplastic GS2 competitively versusl-glutamate, with Ki values in the low micromolar range. At higher concentrations, these inhibitors inactivate glutamine synthetase, while dilution restores activity through enzyme-inhibitor dissociation. Herbicidal phosphinothricins exhibit low Ki values and slow enzyme turnover, as described by reactivation characteristics. Both the GS1 and GS2 isoforms of plant glutamine synthetase are similarly inhibited by the phosphinothricins, consistent with the broad-spectrum herbicidal activity observed for PPT itself as well as other active compounds in this series. PMID:16668090

  7. Weak measurement with orthogonal preselection and postselection

    NASA Astrophysics Data System (ADS)

    Pang, Shengshi; Wu, Shengjun; Chen, Zeng-Bing

    2012-08-01

    Weak measurement is a novel quantum measurement scheme, which is usually characterized by the weak value formalism. To guarantee the validity of the weak value formalism, the fidelity between the preselection and the postselection should not be too small generally. In this work, we study the weak measurement on a qubit system with exactly or asymptotically orthogonal pre- and postselections. We shall establish a general rigorous framework for the weak measurement beyond the weak value formalism, and obtain the average output of a weak measurement when the pre- and postselections are exactly orthogonal. We shall also study the asymptotic behavior of a weak measurement in the limiting process that the pre- and postselections tend to be orthogonal.

  8. Phase array calibration orthogonal phase sequence

    NASA Technical Reports Server (NTRS)

    Sorace, Ronald E. (Inventor); Reinhardt, Victor S. (Inventor); Chan, Clinton (Inventor)

    1999-01-01

    Methods and systems for calibrating an array antenna are described. The array antenna has a plurality of antenna elements each having a signal with a phase and an amplitude forming an array antenna signal. For calibration, the phase of each element signal is sequentially switched one at a time through four orthogonal phase states. At each orthogonal phase state, the power of the array antenna signal is measured. A phase and an amplitude error for each of the element signals is determined based on the power of the array antenna signal at each of the four orthogonal phase states. The phase and amplitude of each of the element signals is then adjusted by the corresponding phase and amplitude errors.

  9. The stable computation of formal orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Beckermann, Bernhard

    1996-12-01

    For many applications - such as the look-ahead variants of the Lanczos algorithm - a sequence of formal (block-)orthogonal polynomials is required. Usually, one generates such a sequence by taking suitable polynomial combinations of a pair of basis polynomials. These basis polynomials are determined by a look-ahead generalization of the classical three term recurrence, where the polynomial coefficients are obtained by solving a small system of linear equations. In finite precision arithmetic, the numerical orthogonality of the polynomials depends on a good choice of the size of the small systems; this size is usually controlled by a heuristic argument such as the condition number of the small matrix of coefficients. However, quite often it happens that orthogonality gets lost.

  10. Orthogonal grid generation in two dimensional space

    NASA Astrophysics Data System (ADS)

    Theodoropoulos, T.; Bergeles, G.; Athanassiadis, N.

    A generalization of a numerical technique for orthogonal mapping, used by Ryskin and Leal (1983) for the construction of boundary-fitted curvilinear coordinate systems in two-dimensional space, is proposed. The boundary-fitted orthogonal curvilinear coordinates are assumed to transform to Cartesian coordinates by Laplace equations. The scale factors involved in the Laplace equations are computed on boundaries and estimated on internal points by means of an interpolation formula. Three types of boundary conditions have been tested: Dirichlet, Cauchy-Riemann, and pseudo-Dirichlet. It is shown that, using this method, grids appropriate for the calculation of flow fields over sharp edges, complex boundary shapes, etc., can be easily constructed. Examples on various geometries are presented, together with a convenient method to check the orthogonality of the resulting meshes.

  11. Recurrent Isolated Neonatal Hemolytic Anemia: Think About Glutathione Synthetase Deficiency.

    PubMed

    Signolet, Isabelle; Chenouard, Rachel; Oca, Florine; Barth, Magalie; Reynier, Pascal; Denis, Marie-Christine; Simard, Gilles

    2016-09-01

    Hemolytic anemia (HA) of the newborn should be considered in cases of rapidly developing, severe, or persistent hyperbilirubinemia. Several causes of corpuscular hemolysis have been described, among which red blood cell enzyme defects are of particular concern. We report a rare case of red blood cell enzyme defect in a male infant, who presented during his first months of life with recurrent and isolated neonatal hemolysis. All main causes were ruled out. At 6.5 months of age, the patient presented with gastroenteritis requiring hospitalization; fortuitously, urine organic acid chromatography revealed a large peak of 5-oxoproline. Before the association between HA and 5-oxoprolinuria was noted, glutathione synthetase deficiency was suspected and confirmed by a low glutathione synthetase concentration and a collapse of glutathione synthetase activity in erythrocytes. Moreover, molecular diagnosis revealed 2 mutations in the glutathione synthetase gene: a previously reported missense mutation (c.[656A>G]; p.[Asp219Gly]) and a mutation not yet described in the binding site of the enzyme (c.[902T>C]; p.[Leu301Pro]). However, 15 days later, a control sample revealed no signs of 5-oxoprolinuria and the clinical history discovered administration of acetaminophen in the 48 hours before hospitalization. Thus, in this patient, acetaminophen exposure allowed the diagnosis of a mild form of glutathione synthetase deficiency, characterized by isolated HA. Early diagnosis is important because treatment with bicarbonate, vitamins C and E, and elimination of trigger factors are recommended to improve long-term outcomes. Glutathione synthetase deficiency should be screened for in cases of unexplained newborn HA. Copyright © 2016 by the American Academy of Pediatrics.

  12. Trellis Coded Modulation Applied to Orthogonal Signals

    DTIC Science & Technology

    1993-03-01

    fti; the codted’on of InO-lmotoo|,. $end cawn.rt, Ih’ M trtldton e I k a ,n nE t# IO.,.(q *S(OI*ftO or In tftri . W•~dtI• % uq ~~O,, ’Ca rdu~ltit9...ABSTRACT (Maximum 200 Wtr) A method for the design of trellis codes for coherent detection of orthogonal signals in additive white Gaussian noise (AWGN...capacity, it is shown that a coding method requiring the same bandwidth efficiency for the orthogonal signal space and maximum likelihood (ML) soft decoding

  13. Orthogonal and Non-Orthogonal Tight Binding Parameters for III-V Semiconductors Nitrides

    NASA Astrophysics Data System (ADS)

    Martins, A. S.; Fellows, C. E.

    2016-12-01

    A simulated annealing (SA) approach is employed in the determination of different tight binding (TB) sets of parameters for the nitride semiconductors AlN, GaN and InN, as well their limitations and potentialities are also discussed. Two kinds of atomic basis set are considered: (i) the orthogonal sp 3 s∗ with interaction up to second neighbors and (ii) a spd non-orthogonal set, with the Hamiltonian matrix elements calculated within the Extended Hückel Theory (EHT) prescriptions. For the non-orthogonal method, TB parameters are given for both zincblend and wurtzite crystalline structures.

  14. Multipartite invariant states. II. Orthogonal symmetry

    SciTech Connect

    Chruscinski, Dariusz; Kossakowski, Andrzej

    2006-06-15

    We construct a class of multipartite states possessing orthogonal symmetry. This new class contains multipartite states which are invariant under the action of local unitary operations introduced in our preceding paper [Phys. Rev. A 73, 062314 (2006)]. We study basic properties of multipartite symmetric states: separability criteria and multi-PPT conditions.

  15. Microcapsules with three orthogonal reactive sites

    PubMed Central

    Mason, Brian P.; Hira, Steven M.; Strouse, Geoffrey F.; McQuade, D. Tyler

    2009-01-01

    Polymeric microcapsules containing reactive sites on the shell surface and two orthogonally reactive polymers encapsulated within the interior are selectively labeled. The capsules provide three spatially separate and differentially reactive sites. Confocal fluorescence microscopy is used to characterize the distribution of labels. Polymers encapsulated are distributed homogeneously within the core and do not interact with the shell even when oppositely charged. PMID:19254010

  16. Simulating Nonequilibrium Radiation via Orthogonal Polynomial Refinement

    DTIC Science & Technology

    2015-01-07

    resolution orthogonal polynomial refinement technique for this multi-disciplinary science. Through the computational mathematics basic research, a...thus the phenomenon must be modeled [1-4]. In addition, the chemical species concentrations and its associated thermodynamic states of an inhomogeneous... thermodynamic state and compositions of the flow medium. The required optical parameters for the nonequilibrium phenomena simulation need to be determined

  17. 2D nearly orthogonal mesh generation

    NASA Astrophysics Data System (ADS)

    Zhang, Yaoxin; Jia, Yafei; Wang, Sam S. Y.

    2004-11-01

    The Ryskin and Leal (RL) system is the most widely used mesh generation system for the orthogonal mapping. However, when this system is used in domains with complex geometry, particularly in those with sharp corners and strong curvatures, serious distortion or overlapping of mesh lines may occur and an acceptable solution may not be possible. In the present study, two methods are proposed to generate nearly orthogonal meshes with the smoothness control. In the first method, the original RL system is modified by introducing smoothness control functions, which are formulated through the blending of the conformal mapping and the orthogonal mapping; while in the second method, the RL system is modified by introducing the contribution factors. A hybrid system of both methods is also developed. The proposed methods are illustrated by several test examples. Applications of these methods in a natural river channel are demonstrated. It is shown that the modified RL systems are capable of producing meshes with an adequate balance between the orthogonality and the smoothness for complex computational domains without mesh distortions and overlapping.

  18. Gram-Schmidt Orthogonalization by Gauss Elimination.

    ERIC Educational Resources Information Center

    Pursell, Lyle; Trimble, S. Y.

    1991-01-01

    Described is the hand-calculation method for the orthogonalization of a given set of vectors through the integration of Gaussian elimination with existing algorithms. Although not numerically preferable, this method adds increased precision as well as organization to the solution process. (JJK)

  19. About orthogonality one system of functions

    NASA Astrophysics Data System (ADS)

    Barmenkov, A. N.

    2017-01-01

    For the modeling of physical processes is proposed to use representation of functions by a special trigonometric system. We present a method for constructing full orthogonal sequences of this type depending on the parameter. This parameter can be selected optimally based on specification of modeling task.

  20. Families of Ellipses and their Orthogonal Trajectories

    ERIC Educational Resources Information Center

    Ayoub, Ayoub B.

    2004-01-01

    The topic of orthogonal trajectories is taught as a geometric application of first order differential equations. Instructors usually elaborate on the concept of a family of curves to emphasize that they are different even if their members are of the same type. In this article the author considers five families of ellipses, discusses their…

  1. Local copying of orthogonal entangled quantum states

    NASA Astrophysics Data System (ADS)

    Anselmi, Fabio; Chefles, Anthony; Plenio, Martin B.

    2004-11-01

    In classical information theory one can, in principle, produce a perfect copy of any input state. In quantum information theory, the no cloning theorem prohibits exact copying of non-orthogonal states. Moreover, if we wish to copy multiparticle entangled states and can perform only local operations and classical communication (LOCC), then further restrictions apply. We investigate the problem of copying orthogonal, entangled quantum states with an entangled blank state under the restriction to LOCC. Throughout, the subsystems have finite dimension D. We show that if all of the states to be copied are non-maximally entangled, then novel LOCC copying procedures based on entanglement catalysis are possible. We then study in detail the LOCC copying problem where both the blank state and at least one of the states to be copied are maximally entangled. For this to be possible, we find that all the states to be copied must be maximally entangled. We obtain a necessary and sufficient condition for LOCC copying under these conditions. For two orthogonal, maximally entangled states, we provide the general solution to this condition. We use it to show that for D = 2, 3, any pair of orthogonal, maximally entangled states can be locally copied using a maximally entangled blank state. However, we also show that for any D which is not prime, one can construct pairs of such states for which this is impossible.

  2. Gram-Schmidt Orthogonalization by Gauss Elimination.

    ERIC Educational Resources Information Center

    Pursell, Lyle; Trimble, S. Y.

    1991-01-01

    Described is the hand-calculation method for the orthogonalization of a given set of vectors through the integration of Gaussian elimination with existing algorithms. Although not numerically preferable, this method adds increased precision as well as organization to the solution process. (JJK)

  3. ABJM amplitudes and the positive orthogonal Grassmannian

    NASA Astrophysics Data System (ADS)

    Huang, Yu-tin; Wen, CongKao

    2014-02-01

    A remarkable connection between perturbative scattering amplitudes of four dimensional planar SYM, and the stratification of the positive Grassmannian, was revealed in the seminal work of Arkani-Hamed et al. Similar extension for three-dimensional ABJM theory was proposed. Here we establish a direct connection between planar scattering amplitudes of ABJM theory, and singularities thereof, to the stratification of the positive orthogonal Grassmannian. In particular, scattering processes are constructed through on-shell diagrams, which are simply iterative gluing of the fundamental four-point amplitude. Each diagram is then equivalent to the merging of fundamental OG2 orthogonal Grassmannian to form a larger OG k , where 2 k is the number of external particles. The invariant information that is encoded in each diagram is precisely this stratification. This information can be easily read off via permutation paths of the on-shell diagram, which also can be used to derive a canonical representation of OG k that manifests the vanishing of consecutive minors as the singularity of all on-shell diagrams. Quite remarkably, for the BCFW recursion representation of the tree-level amplitudes, the on-shell diagram manifests the presence of all physical factorization poles, as well as the cancellation of the spurious poles. After analytically continuing the orthogonal Grassmannian to split signature, we reveal that each on-shell diagram in fact resides in the positive cell of the orthogonal Grassmannian, where all minors are positive. In this language, the amplitudes of ABJM theory is simply an integral of a product of d log forms, over the positive orthogonal Grassmannian.

  4. The Puzzle of Ligand Binding to Corynebacterium ammoniagenes FAD Synthetase*

    PubMed Central

    Frago, Susana; Velázquez-Campoy, Adrián; Medina, Milagros

    2009-01-01

    In bacteria, riboflavin phosphorylation and subsequent conversion of FMN into FAD are carried out by FAD synthetase, a single bifunctional enzyme. Both reactions require ATP and Mg2+. The N-terminal domain of FAD synthetase appears to be responsible for the adenylyltransferase activity, whereas the C-terminal domain would be in charge of the kinase activity. Binding to Corynebacterium ammoniagenes FAD synthetase of its products and substrates, as well as of several analogues, is analyzed. Binding parameters for adenine nucleotides to each one of the two adenine nucleotide sites are reported. In addition, it is demonstrated for the first time that the enzyme presents two independent flavin sites, each one related with one of the enzymatic activities. The binding parameters of flavins to these sites are also provided. The presence of Mg2+ and of both adenine nucleotides and flavins cooperatively modulates the interaction parameters for the other ligands. Our data also suggest that during its double catalytic cycle FAD synthetase must suffer conformational changes induced by adenine nucleotide-Mg2+ or flavin binding. They might include not only rearrangement of the different protein loops but also alternative conformations between domains. PMID:19136717

  5. Polyspecific pyrrolysyl-tRNA synthetases from directed evolution.

    PubMed

    Guo, Li-Tao; Wang, Yane-Shih; Nakamura, Akiyoshi; Eiler, Daniel; Kavran, Jennifer M; Wong, Margaret; Kiessling, Laura L; Steitz, Thomas A; O'Donoghue, Patrick; Söll, Dieter

    2014-11-25

    Pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNA(Pyl) have emerged as ideal translation components for genetic code innovation. Variants of the enzyme facilitate the incorporation >100 noncanonical amino acids (ncAAs) into proteins. PylRS variants were previously selected to acylate N(ε)-acetyl-Lys (AcK) onto tRNA(Pyl). Here, we examine an N(ε)-acetyl-lysyl-tRNA synthetase (AcKRS), which is polyspecific (i.e., active with a broad range of ncAAs) and 30-fold more efficient with Phe derivatives than it is with AcK. Structural and biochemical data reveal the molecular basis of polyspecificity in AcKRS and in a PylRS variant [iodo-phenylalanyl-tRNA synthetase (IFRS)] that displays both enhanced activity and substrate promiscuity over a chemical library of 313 ncAAs. IFRS, a product of directed evolution, has distinct binding modes for different ncAAs. These data indicate that in vivo selections do not produce optimally specific tRNA synthetases and suggest that translation fidelity will become an increasingly dominant factor in expanding the genetic code far beyond 20 amino acids.

  6. Polyspecific pyrrolysyl-tRNA synthetases from directed evolution

    PubMed Central

    Guo, Li-Tao; Wang, Yane-Shih; Nakamura, Akiyoshi; Eiler, Daniel; Kavran, Jennifer M.; Wong, Margaret; Kiessling, Laura L.; Steitz, Thomas A.; O’Donoghue, Patrick; Söll, Dieter

    2014-01-01

    Pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNAPyl have emerged as ideal translation components for genetic code innovation. Variants of the enzyme facilitate the incorporation >100 noncanonical amino acids (ncAAs) into proteins. PylRS variants were previously selected to acylate Nε-acetyl-Lys (AcK) onto tRNAPyl. Here, we examine an Nε-acetyl-lysyl-tRNA synthetase (AcKRS), which is polyspecific (i.e., active with a broad range of ncAAs) and 30-fold more efficient with Phe derivatives than it is with AcK. Structural and biochemical data reveal the molecular basis of polyspecificity in AcKRS and in a PylRS variant [iodo-phenylalanyl-tRNA synthetase (IFRS)] that displays both enhanced activity and substrate promiscuity over a chemical library of 313 ncAAs. IFRS, a product of directed evolution, has distinct binding modes for different ncAAs. These data indicate that in vivo selections do not produce optimally specific tRNA synthetases and suggest that translation fidelity will become an increasingly dominant factor in expanding the genetic code far beyond 20 amino acids. PMID:25385624

  7. A connection between orthogonal polynomials on the unit circle and matrix orthogonal polynomials on the real line

    NASA Astrophysics Data System (ADS)

    Cantero, M. J.; Ferrer, M. P.; Moral, L.; Velázquez, L.

    2003-05-01

    Szego's procedure to connect orthogonal polynomials on the unit circle and orthogonal polynomials on [-1,1] is generalized to nonsymmetric measures. It generates the so-called semi-orthogonal functions on the linear space of Laurent polynomials [Lambda], and leads to a new orthogonality structure in the module [Lambda]×[Lambda]. This structure can be interpreted in terms of a 2×2 matrix measure on [-1,1], and semi-orthogonal functions provide the corresponding sequence of orthogonal matrix polynomials. This gives a connection between orthogonal polynomials on the unit circle and certain classes of matrix orthogonal polynomials on [-1,1]. As an application, the strong asymptotics of these matrix orthogonal polynomials is derived, obtaining an explicit expression for the corresponding Szego's matrix function.

  8. The importance of cytosolic glutamine synthetase in nitrogen assimilation and recycling.

    PubMed

    Bernard, Stéphanie M; Habash, Dimah Z

    2009-01-01

    Glutamine synthetase assimilates ammonium into amino acids, thus it is a key enzyme for nitrogen metabolism. The cytosolic isoenzymes of glutamine synthetase assimilate ammonium derived from primary nitrogen uptake and from various internal nitrogen recycling pathways. In this way, cytosolic glutamine synthetase is crucial for the remobilization of protein-derived nitrogen. Cytosolic glutamine synthetase is encoded by a small family of genes that are well conserved across plant species. Members of the cytosolic glutamine synthetase gene family are regulated in response to plant nitrogen status, as well as to environmental cues, such as nitrogen availability and biotic/abiotic stresses. The complex regulation of cytosolic glutamine synthetase at the transcriptional to post-translational levels is key to the establishment of a specific physiological role for each isoenzyme. The diverse physiological roles of cytosolic glutamine synthetase isoenzymes are important in relation to current agricultural and ecological issues.

  9. Decreased Red Cell Uroporphyrinogen I Synthetase Activity in Intermittent Acute Porphyria

    PubMed Central

    Strand, L. James; Meyer, Urs A.; Felsher, Bertram F.; Redeker, Allan G.; Marver, Harvey S.

    1972-01-01

    Intermittent acute porphyria has recently been distinguished biochemically from other genetic hepatic porphyrias by the observation of diminished hepatic uroporphyrinogen I synthetase activity and increased δ-aminolevulinic acid synthetase activity. Since deficient uroporphyrinogen I synthetase may be reflected in nonhepatic tissues, we have assayed this enzyme in red cell hemolysates from nonporphyric subjects and from patients with genetic hepatic porphyria. Only patients with intermittent acute porphyria had decreased erythrocyte uroporphyrinogen I synthetase activity which was approximately 50% of normal. The apparent Km of partially purified uroporphyrinogen I synthetase was 6 × 10−6m in both nonporphyrics and patients with intermittent acute porphyria. These data provide further evidence for a primary mutation affecting uroporphyrinogen I synthetase in intermittent acute porphyria. Further-more, results of assay of red cell uroporphyrinogen I synthetase activity in a large family with intermittent acute porphyria suggest that this test may be a reliable indicator of the heterozygous state. PMID:5056653

  10. The importance of cytosolic glutamine synthetase in nitrogen assimilation and recycling

    SciTech Connect

    Bernard, S.M.; Habash, D.Z.

    2009-07-02

    Glutamine synthetase assimilates ammonium into amino acids, thus it is a key enzyme for nitrogen metabolism. The cytosolic isoenzymes of glutamine synthetase assimilate ammonium derived from primary nitrogen uptake and from various internal nitrogen recycling pathways. In this way, cytosolic glutamine synthetase is crucial for the remobilization of protein-derived nitrogen. Cytosolic glutamine synthetase is encoded by a small family of genes that are well conserved across plant species. Members of the cytosolic glutamine synthetase gene family are regulated in response to plant nitrogen status, as well as to environmental cues, such as nitrogen availability and biotic/abiotic stresses. The complex regulation of cytosolic glutamine synthetase at the transcriptional to post-translational levels is key to the establishment of a specific physiological role for each isoenzyme. The diverse physiological roles of cytosolic glutamine synthetase isoenzymes are important in relation to current agricultural and ecological issues.

  11. SS{sub p}G: A strongly orthogonal geminal method with relaxed strong orthogonality

    SciTech Connect

    Cagg, Brett A. Rassolov, Vitaly A.

    2014-10-28

    Strong orthogonality is an important constraint placed on geminal wavefunctions in order to make variational minimization tractable. However, strong orthogonality prevents certain, possibly important, excited configurations from contributing to the ground state description of chemical systems. The presented method lifts strong orthogonality constraint from geminal wavefunction by computing a perturbative-like correction to each geminal independently from the corrections to all other geminals. The method is applied to the Singlet-type Strongly orthogonal Geminals variant of the geminal wavefunction. Comparisons of this new SS{sub p}G method are made to the non-orthogonal AP1roG and the unconstrained Geminal Mean-Field Configuration Interaction method using small atomic and molecular systems. The correction is also compared to Density Matrix Renormalization Group calculations performed on long polyene chains in order to assess its scalability and applicability to large strongly correlated systems. The results of these comparisons demonstrate that although the perturbative correction is small, it may be a necessary first step in the systematic improvement of any strongly orthogonal geminal method.

  12. Generalized Orthogonality Relations and SU(1,1)-Quantum Tomography

    NASA Astrophysics Data System (ADS)

    Carmeli, C.; Cassinelli, G.; Zizzi, F.

    2009-06-01

    We present a mathematically precise derivation of some generalized orthogonality relations for the discrete series representations of SU(1,1). These orthogonality relations are applied to derive tomographical reconstruction formulas. Their physical interpretation is also discussed.

  13. Crystal structure of the archaeal asparagine synthetase: interrelation with aspartyl-tRNA and asparaginyl-tRNA synthetases.

    PubMed

    Blaise, Mickaël; Fréchin, Mathieu; Oliéric, Vincent; Charron, Christophe; Sauter, Claude; Lorber, Bernard; Roy, Hervé; Kern, Daniel

    2011-09-23

    Asparagine synthetase A (AsnA) catalyzes asparagine synthesis using aspartate, ATP, and ammonia as substrates. Asparagine is formed in two steps: the β-carboxylate group of aspartate is first activated by ATP to form an aminoacyl-AMP before its amidation by a nucleophilic attack with an ammonium ion. Interestingly, this mechanism of amino acid activation resembles that used by aminoacyl-tRNA synthetases, which first activate the α-carboxylate group of the amino acid to form also an aminoacyl-AMP before they transfer the activated amino acid onto the cognate tRNA. In a previous investigation, we have shown that the open reading frame of Pyrococcus abyssi annotated as asparaginyl-tRNA synthetase (AsnRS) 2 is, in fact, an archaeal asparagine synthetase A (AS-AR) that evolved from an ancestral aspartyl-tRNA synthetase (AspRS). We present here the crystal structure of this AS-AR. The fold of this protein is similar to that of bacterial AsnA and resembles the catalytic cores of AspRS and AsnRS. The high-resolution structures of AS-AR associated with its substrates and end-products help to understand the reaction mechanism of asparagine formation and release. A comparison of the catalytic core of AS-AR with those of archaeal AspRS and AsnRS and with that of bacterial AsnA reveals a strong conservation. This study uncovers how the active site of the ancestral AspRS rearranged throughout evolution to transform an enzyme activating the α-carboxylate group into an enzyme that is able to activate the β-carboxylate group of aspartate, which can react with ammonia instead of tRNA. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Clone and functional analysis of Seryl-tRNA synthetase and Tyrosyl-tRNA synthetase from silkworm, Bombyx mori.

    PubMed

    Hu, Jingsheng; Tian, Jianghai; Li, Fanchi; Xue, Bin; Hu, Jiahuan; Cheng, Xiaoyu; Li, Jinxin; Shen, Weide; Li, Bing

    2017-01-30

    Aminoacyl-tRNA synthetases are the key enzymes for protein synthesis. Glycine, alanine, serine and tyrosine are the major amino acids composing fibroin of silkworm. Among them, the genes of alanyl-tRNA synthetase (AlaRS) and glycyl-tRNA synthetase (GlyRS) have been cloned. In this study, the seryl-tRNA synthetase (SerRS) and tyrosyl-tRNA synthetase (TyrRS) genes from silkworm were cloned. Their full length are 1709 bp and 1868 bp and contain open reading frame (ORF) of 1485 bp and 1575 bp, respectively. RT-PCR examination showed that the transcription levels of SerRS, TyrRS, AlaRS and GlyRS are significantly higher in silk gland than in other tissues. In addition, their transcription levels are much higher in middle and posterior silk gland than in anterior silk gland. Moreover, treatment of silkworms with phoxim, an inhibitor of silk protein synthesis, but not TiO2 NP, an enhancer of silk protein synthesis, significantly reduced the transcription levels of aaRS and content of free amino acids in posterior silk gland, therefore affecting silk protein synthesis, which may be the mechanism of phoxim-silking disorders. Furthermore, low concentration of TiO2 NPs showed no effect on the transcription of aaRS and content of free amino acids, suggesting that TiO2 NPs promotes silk protein synthesis possibly by increasing the activity of fibroin synthase in silkworm.

  15. Clone and functional analysis of Seryl-tRNA synthetase and Tyrosyl-tRNA synthetase from silkworm, Bombyx mori

    PubMed Central

    Hu, Jingsheng; Tian, Jianghai; Li, Fanchi; Xue, Bin; Hu, Jiahuan; Cheng, Xiaoyu; Li, Jinxin; Shen, Weide; Li, Bing

    2017-01-01

    Aminoacyl-tRNA synthetases are the key enzymes for protein synthesis. Glycine, alanine, serine and tyrosine are the major amino acids composing fibroin of silkworm. Among them, the genes of alanyl-tRNA synthetase (AlaRS) and glycyl-tRNA synthetase (GlyRS) have been cloned. In this study, the seryl-tRNA synthetase (SerRS) and tyrosyl-tRNA synthetase (TyrRS) genes from silkworm were cloned. Their full length are 1709 bp and 1868 bp and contain open reading frame (ORF) of 1485 bp and 1575 bp, respectively. RT-PCR examination showed that the transcription levels of SerRS, TyrRS, AlaRS and GlyRS are significantly higher in silk gland than in other tissues. In addition, their transcription levels are much higher in middle and posterior silk gland than in anterior silk gland. Moreover, treatment of silkworms with phoxim, an inhibitor of silk protein synthesis, but not TiO2 NP, an enhancer of silk protein synthesis, significantly reduced the transcription levels of aaRS and content of free amino acids in posterior silk gland, therefore affecting silk protein synthesis, which may be the mechanism of phoxim-silking disorders. Furthermore, low concentration of TiO2 NPs showed no effect on the transcription of aaRS and content of free amino acids, suggesting that TiO2 NPs promotes silk protein synthesis possibly by increasing the activity of fibroin synthase in silkworm. PMID:28134300

  16. Weak lensing tomography with orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Schäfer, Björn Malte; Heisenberg, Lavinia

    2012-07-01

    The topic of this paper is weak cosmic shear tomography where the line-of-sight weighting is carried out with a set of specifically constructed orthogonal polynomials, dubbed Tomography with Orthogonal Radial Distance Polynomial Systems (TaRDiS). We investigate the properties of these polynomials and employ weak convergence spectra, which have been obtained by weighting with these polynomials, for the estimation of cosmological parameters. We quantify their power in constraining parameters in a Fisher matrix technique and demonstrate how each polynomial projects out statistically independent information, and how the combination of multiple polynomials lifts degeneracies. The assumption of a reference cosmology is needed for the construction of the polynomials, and as a last point we investigate how errors in the construction with a wrong cosmological model propagate to misestimates in cosmological parameters. TaRDiS performs on a similar level as traditional tomographic methods and some key features of tomography are made easier to understand.

  17. Greedy Criterion in Orthogonal Greedy Learning.

    PubMed

    Xu, Lin; Lin, Shaobo; Zeng, Jinshan; Liu, Xia; Fang, Yi; Xu, Zongben

    2017-02-23

    Orthogonal greedy learning (OGL) is a stepwise learning scheme that starts with selecting a new atom from a specified dictionary via the steepest gradient descent (SGD) and then builds the estimator through orthogonal projection. In this paper, we found that SGD is not the unique greedy criterion and introduced a new greedy criterion, called as ''δ-greedy threshold'' for learning. Based on this new greedy criterion, we derived a straightforward termination rule for OGL. Our theoretical study shows that the new learning scheme can achieve the existing (almost) optimal learning rate of OGL. Numerical experiments are also provided to support that this new scheme can achieve almost optimal generalization performance while requiring less computation than OGL.

  18. Orthogonal photoswitching in a multifunctional molecular system

    PubMed Central

    Lerch, Michael M.; Hansen, Mickel J.; Velema, Willem A.; Szymanski, Wiktor; Feringa, Ben L.

    2016-01-01

    The wavelength-selective, reversible photocontrol over various molecular processes in parallel remains an unsolved challenge. Overlapping ultraviolet-visible spectra of frequently employed photoswitches have prevented the development of orthogonally responsive systems, analogous to those that rely on wavelength-selective cleavage of photo-removable protecting groups. Here we report the orthogonal and reversible control of two distinct types of photoswitches in one solution, that is, a donor–acceptor Stenhouse adduct (DASA) and an azobenzene. The control is achieved by using three different wavelengths of irradiation and a thermal relaxation process. The reported combination tolerates a broad variety of differently substituted photoswitches. The presented system is also extended to an intramolecular combination of photoresponsive units. A model application for an intramolecular combination of switches is presented, in which the DASA component acts as a phase-transfer tag, while the azobenzene moiety independently controls the binding to α-cyclodextrin. PMID:27401266

  19. An orientation-selective orthogonal lapped transform.

    PubMed

    Kunz, Dietmar

    2008-08-01

    A novel critically sampled orientation-selective orthogonal lapped transform called the lapped Hartley transform (LHT) is derived. In a first step, overlapping basis functions are generated by modulating basis functions of a 2-D block Hartley transform by a cosine wave. To achieve invertibility and orthogonality, an iterative filter is applied as prefilter in the analysis and as postfilter in the synthesis operation, respectively. Alternatively, filtering can be restricted to analysis or synthesis, ending up with a biorthogonal transform (LHT-PR, LHT-PO). A statistical analysis based on a 4000-image data base shows that the LHT and LHT-PO have better redundancy removal properties than other block or lapped transforms. Finally, image compression and noise removal examples are given, showing the advantages of the LHT especially in images containing oriented textures.

  20. HOLA: Human-like Orthogonal Network Layout.

    PubMed

    Kieffer, Steve; Dwyer, Tim; Marriott, Kim; Wybrow, Michael

    2016-01-01

    Over the last 50 years a wide variety of automatic network layout algorithms have been developed. Some are fast heuristic techniques suitable for networks with hundreds of thousands of nodes while others are multi-stage frameworks for higher-quality layout of smaller networks. However, despite decades of research currently no algorithm produces layout of comparable quality to that of a human. We give a new "human-centred" methodology for automatic network layout algorithm design that is intended to overcome this deficiency. User studies are first used to identify the aesthetic criteria algorithms should encode, then an algorithm is developed that is informed by these criteria and finally, a follow-up study evaluates the algorithm output. We have used this new methodology to develop an automatic orthogonal network layout method, HOLA, that achieves measurably better (by user study) layout than the best available orthogonal layout algorithm and which produces layouts of comparable quality to those produced by hand.

  1. Multicarrier orthogonal spread-spectrum (MOSS) data communications

    DOEpatents

    Smith, Stephen F.; Dress, William B.

    2008-01-01

    Systems and methods are described for multicarrier orthogonal spread-spectrum (MOSS) data communication. A method includes individually spread-spectrum modulating at least two of a set of orthogonal frequency division multiplexed carriers, wherein the resulting individually spread-spectrum modulated at least two of a set of orthogonal frequency division multiplexed carriers are substantially mutually orthogonal with respect to both frequency division multiplexing and spread-spectrum modulation.

  2. Complete spacelike hypersurfaces in orthogonally splitted spacetimes

    NASA Astrophysics Data System (ADS)

    Colombo, Giulio; Rigoli, Marco

    2017-10-01

    We provide some "half-space theorems" for spacelike complete non-compact hypersurfaces into orthogonally splitted spacetimes. In particular we generalize some recent work of Rubio and Salamanca on maximal spacelike compact hypersurfaces. Beside compactness, we also relax some of their curvature assumptions and even consider the case of nonconstant mean curvature bounded from above. The analytic tools used in various arguments are based on some forms of the weak maximum principle.

  3. Parallel Spectral Acquisition with Orthogonal ICR Cells.

    PubMed

    Park, Sung-Gun; Anderson, Gordon A; Bruce, James E

    2017-03-01

    FT-based high performance mass analyzers yield increased resolving power and mass measurement accuracy, yet require increased duration of signal acquisition that can limit many applications. The implementation of stronger magnetic fields, multiple detection electrodes for harmonic signal detection, and an array of multiple mass analyzers arranged along the magnetic field axis have been used to decrease required acquisition time. The results presented here show that multiple ion cyclotron resonance (ICR) mass analyzers can also be implemented orthogonal to the central magnetic field axis. The orthogonal ICR cell system presented here consisting of two cells (master and slave cells) was constructed with printed circuit boards and installed within a single superconducting magnet and vacuum system. A master cell was positioned, as is normally done with ICR cells, on the central magnetic field axis and a slave cell was located off this central axis, but directly adjacent and alongside the master cell. To achieve ion transfer between cells, ions that were initially trapped in the master cell were drifted across the magnetic field into the slave cell with application of a small DC field applied perpendicularly to the magnetic field axis. A subsequent population of ions was injected and accumulated in the master cell. Simultaneous excitation of cyclotron motion of ions in both cells was carried out; ICR signals from each cell were independently amplified and recorded in parallel. Presented here are the initial results of successful parallel spectral acquisition with this orthogonal dual ICR cell array. Graphical Abstract ᅟ.

  4. Parallel Spectral Acquisition with Orthogonal ICR Cells

    NASA Astrophysics Data System (ADS)

    Park, Sung-Gun; Anderson, Gordon A.; Bruce, James E.

    2017-03-01

    FT-based high performance mass analyzers yield increased resolving power and mass measurement accuracy, yet require increased duration of signal acquisition that can limit many applications. The implementation of stronger magnetic fields, multiple detection electrodes for harmonic signal detection, and an array of multiple mass analyzers arranged along the magnetic field axis have been used to decrease required acquisition time. The results presented here show that multiple ion cyclotron resonance (ICR) mass analyzers can also be implemented orthogonal to the central magnetic field axis. The orthogonal ICR cell system presented here consisting of two cells (master and slave cells) was constructed with printed circuit boards and installed within a single superconducting magnet and vacuum system. A master cell was positioned, as is normally done with ICR cells, on the central magnetic field axis and a slave cell was located off this central axis, but directly adjacent and alongside the master cell. To achieve ion transfer between cells, ions that were initially trapped in the master cell were drifted across the magnetic field into the slave cell with application of a small DC field applied perpendicularly to the magnetic field axis. A subsequent population of ions was injected and accumulated in the master cell. Simultaneous excitation of cyclotron motion of ions in both cells was carried out; ICR signals from each cell were independently amplified and recorded in parallel. Presented here are the initial results of successful parallel spectral acquisition with this orthogonal dual ICR cell array.

  5. Orthogonal ion injection apparatus and process

    DOEpatents

    Kurulugama, Ruwan T; Belov, Mikhail E

    2014-04-15

    An orthogonal ion injection apparatus and process are described in which ions are directly injected into an ion guide orthogonal to the ion guide axis through an inlet opening located on a side of the ion guide. The end of the heated capillary is placed inside the ion guide such that the ions are directly injected into DC and RF fields inside the ion guide, which efficiently confines ions inside the ion guide. Liquid droplets created by the ionization source that are carried through the capillary into the ion guide are removed from the ion guide by a strong directional gas flow through an inlet opening on the opposite side of the ion guide. Strong DC and RF fields divert ions into the ion guide. In-guide orthogonal injection yields a noise level that is a factor of 1.5 to 2 lower than conventional inline injection known in the art. Signal intensities for low m/z ions are greater compared to convention inline injection under the same processing conditions.

  6. Direct energy functional minimization under orthogonality constraints

    NASA Astrophysics Data System (ADS)

    Weber, Valéry; VandeVondele, Joost; Hutter, Jürg; Niklasson, Anders M. N.

    2008-02-01

    The direct energy functional minimization problem in electronic structure theory, where the single-particle orbitals are optimized under the constraint of orthogonality, is explored. We present an orbital transformation based on an efficient expansion of the inverse factorization of the overlap matrix that keeps orbitals orthonormal. The orbital transformation maps the orthogonality constrained energy functional to an approximate unconstrained functional, which is correct to some order in a neighborhood of an orthogonal but approximate solution. A conjugate gradient scheme can then be used to find the ground state orbitals from the minimization of a sequence of transformed unconstrained electronic energy functionals. The technique provides an efficient, robust, and numerically stable approach to direct total energy minimization in first principles electronic structure theory based on tight-binding, Hartree-Fock, or density functional theory. For sparse problems, where both the orbitals and the effective single-particle Hamiltonians have sparse matrix representations, the effort scales linearly with the number of basis functions N in each iteration. For problems where only the overlap and Hamiltonian matrices are sparse the computational cost scales as O(M2N ), where M is the number of occupied orbitals. We report a single point density functional energy calculation of a DNA decamer hydrated with 4003 water molecules under periodic boundary conditions. The DNA fragment containing a cis-syn thymine dimer is composed of 634 atoms and the whole system contains a total of 12 661 atoms and 103 333 spherical Gaussian basis functions.

  7. Parallel Spectral Acquisition with Orthogonal ICR Cells

    NASA Astrophysics Data System (ADS)

    Park, Sung-Gun; Anderson, Gordon A.; Bruce, James E.

    2017-01-01

    FT-based high performance mass analyzers yield increased resolving power and mass measurement accuracy, yet require increased duration of signal acquisition that can limit many applications. The implementation of stronger magnetic fields, multiple detection electrodes for harmonic signal detection, and an array of multiple mass analyzers arranged along the magnetic field axis have been used to decrease required acquisition time. The results presented here show that multiple ion cyclotron resonance (ICR) mass analyzers can also be implemented orthogonal to the central magnetic field axis. The orthogonal ICR cell system presented here consisting of two cells (master and slave cells) was constructed with printed circuit boards and installed within a single superconducting magnet and vacuum system. A master cell was positioned, as is normally done with ICR cells, on the central magnetic field axis and a slave cell was located off this central axis, but directly adjacent and alongside the master cell. To achieve ion transfer between cells, ions that were initially trapped in the master cell were drifted across the magnetic field into the slave cell with application of a small DC field applied perpendicularly to the magnetic field axis. A subsequent population of ions was injected and accumulated in the master cell. Simultaneous excitation of cyclotron motion of ions in both cells was carried out; ICR signals from each cell were independently amplified and recorded in parallel. Presented here are the initial results of successful parallel spectral acquisition with this orthogonal dual ICR cell array.

  8. Essentiality Assessment of Cysteinyl and Lysyl-tRNA Synthetases of Mycobacterium smegmatis

    PubMed Central

    Ravishankar, Sudha; Ambady, Anisha; Swetha, Rayapadi G.; Anbarasu, Anand; Ramaiah, Sudha; Sambandamurthy, Vasan K.

    2016-01-01

    Discovery of mupirocin, an antibiotic that targets isoleucyl-tRNA synthetase, established aminoacyl-tRNA synthetase as an attractive target for the discovery of novel antibacterial agents. Despite a high degree of similarity between the bacterial and human aminoacyl-tRNA synthetases, the selectivity observed with mupirocin triggered the possibility of targeting other aminoacyl-tRNA synthetases as potential drug targets. These enzymes catalyse the condensation of a specific amino acid to its cognate tRNA in an energy-dependent reaction. Therefore, each organism is expected to encode at least twenty aminoacyl-tRNA synthetases, one for each amino acid. However, a bioinformatics search for genes encoding aminoacyl-tRNA synthetases from Mycobacterium smegmatis returned multiple genes for glutamyl (GluRS), cysteinyl (CysRS), prolyl (ProRS) and lysyl (LysRS) tRNA synthetases. The pathogenic mycobacteria, namely, Mycobacterium tuberculosis and Mycobacterium leprae, were also found to possess two genes each for CysRS and LysRS. A similar search indicated the presence of additional genes for LysRS in gram negative bacteria as well. Herein, we describe sequence and structural analysis of the additional aminoacyl-tRNA synthetase genes found in M. smegmatis. Characterization of conditional expression strains of Cysteinyl and Lysyl-tRNA synthetases generated in M. smegmatis revealed that the canonical aminoacyl-tRNA synthetase are essential, while the additional ones are not essential for the growth of M. smegmatis. PMID:26794499

  9. Inhibition of recombinant Pneumocystis carinii dihydropteroate synthetase by sulfa drugs.

    PubMed

    Hong, Y L; Hossler, P A; Calhoun, D H; Meshnick, S R

    1995-08-01

    Forty-four sulfa drugs were screened against crude preparations of recombinant Pneumocystis carinii dihydropteroate synthetase. The apparent Michaelis-Menten constants (Km) for p-aminobenzoic acid and 7,8-dihydro-6-hydroxymethylpterin pyrophosphate were 0.34 +/- 0.02 and 2.50 +/- 0.71 microM, respectively. Several sulfa drugs, including sulfathiazole, sulfachlorpyridazine, sulfamethoxypyridazine, and sulfathiourea, inhibited dihydropteroate synthetase approximately as well as sulfamethoxazole, as determined by the concentrations which cause 50% inhibition and/or by Ki. For all sulfones and sulfonamides tested, unsubstituted p-amino groups were necessary for activity, and sulfonamides containing an N1-heterocyclic substituent were found to be the most effective inhibitors. Folate biosynthesis in isolated intact P. carinii was approximately equally sensitive to inhibition by sulfamethoxazole, sulfachlorpyridazine, sulfamethoxypyridazine, sulfisoxazole, and sulfathiazole. Two of these drugs, sulfamethoxypyridazine and sulfisoxazole, are known to be less toxic than sulfamethoxazole and should be further evaluated for the treatment of P. carinii pneumonia.

  10. Aminoacyl-tRNA Synthetase Complexes in Evolution

    PubMed Central

    Havrylenko, Svitlana; Mirande, Marc

    2015-01-01

    Aminoacyl-tRNA synthetases are essential enzymes for interpreting the genetic code. They are responsible for the proper pairing of codons on mRNA with amino acids. In addition to this canonical, translational function, they are also involved in the control of many cellular pathways essential for the maintenance of cellular homeostasis. Association of several of these enzymes within supramolecular assemblies is a key feature of organization of the translation apparatus in eukaryotes. It could be a means to control their oscillation between translational functions, when associated within a multi-aminoacyl-tRNA synthetase complex (MARS), and nontranslational functions, after dissociation from the MARS and association with other partners. In this review, we summarize the composition of the different MARS described from archaea to mammals, the mode of assembly of these complexes, and their roles in maintenance of cellular homeostasis. PMID:25807264

  11. Biochemical and inhibition studies of glutamine synthetase from Leishmania donovani.

    PubMed

    Kumar, Vinay; Yadav, Shailendra; Soumya, Neelagiri; Kumar, Rohit; Babu, Neerupudi Kishore; Singh, Sushma

    2017-03-25

    Leishmaniasis is a group of tropical diseases caused by protozoan parasites of the genus Leishmania. Leishmania donovani is a protozoan parasite that causes visceral leishmaniasis, a fatal disease if left untreated. Chemotherapy for leishmaniasis is problematic as the available drugs are toxic, costly and shows drug resistance, hence, there is a necessity to look out for the novel drug targets, chemical entities and vaccine. Glutamine synthetase (GS) catalyzes the synthesis of glutamine from glutamate and ammonia. In the present study, we have identified and characterized GS from L. donovani. The nucleotide sequence encoding putative glutamine synthetase like sequence from L. donovani (LdGS, LDBPK_060370) was cloned. A 43.5 kDa protein with 6X-His tag at the C-terminal end was obtained by overexpression of LdGS in Escherichia coli BL21 (DE3) strain. Expression of native LdGS in promastigotes and recombinant L. donovani glutamine synthetase (rLdGS) was confirmed by western blot analysis. An increase in expression of GS was observed at different phases of growth of the parasite. Expression of LdGS in promastigote and amastigote was confirmed by western blot analysis. Immunofluorescence studies of both the promastigote and amastigote stages of the parasite revealed the presence of LdGS in cytoplasm. GS exists as a single copy gene in parasite genome. Kinetic analysis of GS enzyme revealed Km value of 26.3 ± 0.4 mM for l- glutamate and Vmax value of 2.15 ± 0.07 U mg(-1). Present study confirms the presence of glutamine synthetase in L. donovani and provides comprehensive overview of LdGS for further validating it as a potential drug target.

  12. Positional isotope exchange analysis of the pantothenate synthetase reaction.

    PubMed

    Williams, LaKenya; Zheng, Renjian; Blanchard, John S; Raushel, Frank M

    2003-05-06

    Pantothenate synthetase from Mycobacterium tuberculosis catalyzes the formation of pantothenate from ATP, D-pantoate, and beta-alanine. The formation of a kinetically competent pantoyl-adenylate intermediate was established by the observation of a positional isotope exchange (PIX) reaction within (18)O-labeled ATP in the presence of d-pantoate. When [betagamma-(18)O(6)]-ATP was incubated with pantothenate synthetase in the presence of d-pantoate, an (18)O label gradually appeared in the alphabeta-bridge position from both the beta- and the gamma-nonbridge positions. The rates of these two PIX reactions were followed by (31)P NMR spectroscopy and found to be identical. These results are consistent with the formation of enzyme-bound pantoyl-adenylate and pyrophosphate upon the mixing of ATP, D-pantoate, and enzyme. In addition, these results require the complete torsional scrambling of the two phosphoryl groups of the labeled pyrophosphate product. The rate of the PIX reaction increased as the D-pantoate concentration was elevated and then decreased to zero at saturating levels of D-pantoate. These inhibition results support the ordered binding of ATP and D-pantoate to the enzyme active site. The PIX reaction was abolished with the addition of pyrophosphatase; thus, PP(i) must be free to dissociate from the active site upon formation of the pantoyl-adenylate intermediate. The PIX reaction rate diminished when the concentrations of ATP and D-pantoate were held constant and the concentration of the third substrate, beta-alanine, was increased. This observation is consistent with a kinetic mechanism that requires the binding of beta-alanine after the release of pyrophosphate from the active site of pantothenate synthetase. Positional isotope exchange reactions have therefore demonstrated that pantothenate synthetase catalyzes the formation of a pantoyl-adenylate intermediate upon the ordered addition of ATP and pantoate.

  13. A Rationally Engineered Misacylating Aminoacyl-Trna Synthetase

    SciTech Connect

    Bullock, T.L.; Rodriguez-Hernandez, A.; Corigliano, E.M.; Perona, J.J.

    2009-05-12

    Information transfer from nucleic acid to protein is mediated by aminoacyl-tRNA synthetases, which catalyze the specific pairings of amino acids with transfer RNAs. Despite copious sequence and structural information on the 22 tRNA synthetase families, little is known of the enzyme signatures that specify amino acid selectivities. Here, we show that transplanting a conserved arginine residue from glutamyl-tRNA synthetase (GluRS) to glutaminyl-tRNA synthetase (GlnRS) improves the K{sub M} of GlnRS for noncognate glutamate. Two crystal structures of this C229R GlnRS mutant reveal that a conserved twin-arginine GluRS amino acid identity signature cannot be incorporated into GlnRS without disrupting surrounding protein structural elements that interact with the tRNA. Consistent with these findings, we show that cumulative replacement of other primary binding site residues in GlnRS, with those of GluRS, only slightly improves the ability of the GlnRS active site to accommodate glutamate. However, introduction of 22 amino acid replacements and one deletion, including substitution of the entire primary binding site and two surface loops adjacent to the region disrupted in C229R, improves the capacity of Escherichia coli GlnRS to synthesize misacylated Glu-tRNA{sup Gln} by 16,000-fold. This hybrid enzyme recapitulates the function of misacylating GluRS enzymes found in organisms that synthesize Gln-tRNA{sup Gln} by an alternative pathway. These findings implicate the RNA component of the contemporary GlnRS-tRNA{sup Gln} complex in mediating amino acid specificity. This role for tRNA may persist as a relic of primordial cells in which the evolution of the genetic code was driven by RNA-catalyzed amino acid-RNA pairing.

  14. Identification of the glutamine synthetase adenylyltransferase of Azospirillum brasilense.

    PubMed

    Van Dommelen, Anne; Spaepen, Stijn; Vanderleyden, Jozef

    2009-04-01

    Glutamine synthetase, a key enzyme in nitrogen metabolism of both prokaryotes and eukaryotes, is strictly regulated. One means of regulation is the modulation of activity through adenylylation catalyzed by adenylyltransferases. Using PCR primers based on conserved sequences in glutamine synthetase adenylyltransferases, we amplified part of the glnE gene of Azospirillum brasilense Sp7. The complete glnE sequence of A. brasilense Sp245 was retrieved from the draft genome sequence of this organism (http://genomics.ornl.gov/research/azo/). Adenylyltransferase is a bifunctional enzyme consisting of an N-terminal domain responsible for deadenylylation activity and a C-terminal domain responsible for adenylylation activity. Both domains are partially homologous to each other. Residues important for catalytic activity were present in the deduced amino acid sequence of the A. brasilense Sp245 glnE sequence. A glnE mutant was constructed in A. brasilense Sp7 by inserting a kanamycin resistance cassette between the two active domains of the enzyme. The resulting mutant was unable to adenylylate the glutamine synthetase enzyme and was impaired in growth when shifted from nitrogen-poor to nitrogen-rich medium.

  15. Turnover of bacterial glutamine synthetase: oxidative inactivation precedes proteolysis.

    PubMed

    Levine, R L; Oliver, C N; Fulks, R M; Stadtman, E R

    1981-04-01

    We partially purified a preparation from Escherichia coli that proteolytically degrades the enzyme glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2]. The degradation is at least a two-step process. First, the glutamine synthetase undergoes an oxidative modification. This modification leads to loss of catalytic activity and also renders the protein susceptible to proteolytic attack in the second step. The oxidative step displays characteristics of a mixed-function oxidation, requiring both molecular oxygen and a reduced nucleotide. This step can also be catalyzed by a purified, mammalian cytochrome P-450 system, as well as by a model system consisting of ascorbic acid and oxygen. Catalase blocks this oxidative modification step. Thus, the overall process of proteolytic degradation can be observed only if care is taken to remove catalase activity from the extracts. The inactivation reaction is dependent on the state of adenylylation of the glutamine synthetase, suggesting that this a physiologically important reaction. If so, then mixed-function oxidases are now implicated in the process of intracellular protein turnover.

  16. Turnover of bacterial glutamine synthetase: oxidative inactivation precedes proteolysis.

    PubMed Central

    Levine, R L; Oliver, C N; Fulks, R M; Stadtman, E R

    1981-01-01

    We partially purified a preparation from Escherichia coli that proteolytically degrades the enzyme glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2]. The degradation is at least a two-step process. First, the glutamine synthetase undergoes an oxidative modification. This modification leads to loss of catalytic activity and also renders the protein susceptible to proteolytic attack in the second step. The oxidative step displays characteristics of a mixed-function oxidation, requiring both molecular oxygen and a reduced nucleotide. This step can also be catalyzed by a purified, mammalian cytochrome P-450 system, as well as by a model system consisting of ascorbic acid and oxygen. Catalase blocks this oxidative modification step. Thus, the overall process of proteolytic degradation can be observed only if care is taken to remove catalase activity from the extracts. The inactivation reaction is dependent on the state of adenylylation of the glutamine synthetase, suggesting that this a physiologically important reaction. If so, then mixed-function oxidases are now implicated in the process of intracellular protein turnover. Images PMID:6113590

  17. Mitochondrial aminoacyl-tRNA synthetases in human disease.

    PubMed

    Konovalova, Svetlana; Tyynismaa, Henna

    2013-04-01

    Mitochondrial aminoacyl-tRNA synthetases (mtARSs) are essential in the process of transferring genetic information from mitochondrial DNA to the complexes of the oxidative phosphorylation system. These synthetases perform an integral step in the initiation of mitochondrial protein synthesis by charging tRNAs with their cognate amino acids. All mtARSs are encoded by nuclear genes, nine of which have recently been described as disease genes for mitochondrial disorders. Unexpectedly, the clinical presentations of these diseases are highly specific to the affected synthetase. Encephalopathy is the most common manifestation but again with gene-specific outcomes. Other clinical presentations include myopathy with anemia, cardiomyopathy, tubulopathy and hearing loss with female ovarian dysgenesis. Here we review the described mutation types and the associated patient phenotypes. The identified mutation spectrum suggests that only mutation types that allow some residual tRNA-charging activity can result in the described mtARS diseases but the molecular mechanisms behind the selective tissue involvement are not currently understood.

  18. Expression of glutamine synthetase in balloon cells: a basis of their antiepileptic role?

    PubMed

    Buccoliero, Anna Maria; Barba, Carmen; Giordano, Flavio; Baroni, Gianna; Genitori, Lorenzo; Guerrini, Renzo; Taddei, Gian Luigi

    2015-01-01

    Glutamine synthetase is an enzyme involved in the clearance of glutamate, the most potent excitatory neurotransmitter. We studied the immunohistochemical expression of glutamine synthetase in neocortical samples from 5 children who underwent surgery for pharmacoresistant epilepsy and a histological diagnosis of focal cortical dysplasia IIb. In all cases, balloon cells, but not dysmorphic neurons, were immunopositive for glutamine synthetase. This finding suggests that balloon cells can be involved in the neutralization of glutamate and play a protective anti-seizure role.

  19. Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase

    SciTech Connect

    Bonnefond, Luc; Frugier, Magali; Touzé, Elodie; Lorber, Bernard; Florentz, Catherine; Giegé, Richard Rudinger-Thirion, Joëlle; Sauter, Claude

    2007-04-01

    Crystals of human mitochondrial tyrosyl-tRNA synthetase lacking the C-terminal S4-like domain diffract to 2.7 Å resolution and are suitable for structure determination. Human mitochondrial tyrosyl-tRNA synthetase and a truncated version with its C-terminal S4-like domain deleted were purified and crystallized. Only the truncated version, which is active in tyrosine activation and Escherichia coli tRNA{sup Tyr} charging, yielded crystals suitable for structure determination. These tetragonal crystals, belonging to space group P4{sub 3}2{sub 1}2, were obtained in the presence of PEG 4000 as a crystallizing agent and diffracted X-rays to 2.7 Å resolution. Complete data sets could be collected and led to structure solution by molecular replacement.

  20. Reciprocity and orthogonality relations for ring resonators

    SciTech Connect

    Wright, E.M.; Obrien, D.P.

    1984-12-01

    A general and rigorous derivation of the reciprocity and orthogonality relations for ring resonators is given without resorting to matrix representations. The general form of the integral equations appropriate to the study of ring resonators containing at least one hard aperture is discussed, and a reciprocity relation for a very general class of ring resonator is established using a theorem concerning so-called Hilbert-Schmidt kernels. It is shown that under very general conditions linear ring resonators are reciprocal and that the transverse eigenmodes for propagation in any direction around the resonator are biorthogonal to those for propagation in the opposite direction. 17 references.

  1. Proper orthogonal decomposition of solar photospheric motions.

    PubMed

    Vecchio, A; Carbone, V; Lepreti, F; Primavera, L; Sorriso-Valvo, L; Veltri, P; Alfonsi, G; Straus, Th

    2005-08-05

    The spatiotemporal dynamics of the solar photosphere is studied by performing a proper orthogonal decomposition (POD) of line of sight velocity fields computed from high resolution data coming from the MDI/SOHO instrument. Using this technique, we are able to identify and characterize the different dynamical regimes acting in the system. Low-frequency oscillations, with frequencies in the range 20-130 microHz, dominate the most energetic POD modes (excluding solar rotation), and are characterized by spatial patterns with typical scales of about 3 Mm. Patterns with larger typical scales of approximately 10 Mm, are associated to p-modes oscillations at frequencies of about 3000 microHz.

  2. Tunable RF Cavities Using Orthogonally Biased Ferrite

    SciTech Connect

    Johnson, R.P.; Alsharo'a, M.; Ankenbrandt, C.M.; Entchevitch, I.; Griffin, J.E.; Moretti, A.; Popovic, M.; Tomlin, R.; /Fermilab

    2009-05-01

    Originally conceived as a solution for FFAG applications, a new compact RF cavity design that tunes rapidly over various frequency ranges can be used to upgrade existing machines. The design being developed uses orthogonally biased garnet cores for fast frequency tuning and liquid dielectric to adjust the frequency range and to control the core temperature. We describe measurements of candidate ferrite and dielectric materials. The first use of the new cavity concept will be for improvements to the 8 GeV Fermilab Booster synchrotron.

  3. Orthogonal Synthesis of Xeno Nucleic Acids.

    PubMed

    Fiers, Guillaume; Chouikhi, Dalila; Oswald, Laurence; Al Ouahabi, Abdelaziz; Chan-Seng, Delphine; Charles, Laurence; Lutz, Jean-François

    2016-12-12

    Sequence-defined peptide triazole nucleic acids (PTzNA) were synthesized by means of a solid-phase orthogonal "AB+CD" iterative strategy. In this approach, AB and CD building blocks containing carboxylic acid (A), azide (B), alkyne (C), and primary amine (D) functions are assembled together by successive copper-catalyzed azide-alkyne cycloaddition (CuAAC) and acid-amine coupling steps. Different PTzNA genetic sequences were prepared using a library of eight building blocks (i.e., four AB and four CD building blocks).

  4. Discriminating orthogonal single-photon images

    NASA Astrophysics Data System (ADS)

    Broadbent, Curtis J.; Zerom, Petros; Shin, Heedeuk; Howell, John C.; Boyd, Robert W.

    2009-03-01

    We can encode an image from an orthogonal basis set onto a single photon from a downconverted pair via the use of an amplitude mask. We can then discriminate the image imprinted on the photon from other images in the set using holographic-matched filtering techniques. We demonstrate this procedure experimentally for an image space of two objects, and we discuss the possibility of applying this method to a much larger image space. This process could have important implications for the manipulation of images at the quantum level.

  5. Intensity correlation of orthogonally crossed speckle waves

    NASA Astrophysics Data System (ADS)

    Okamoto, Takashi

    2006-05-01

    The statistical properties of three-dimensional laser speckles formed with orthogonally crossed multiple speckle waves are studied theoretically. The analytical expressions for the first and second order statistics of the crossed speckle fields are derived, and the theoretical results are verified by means of computer simulations. In the simulations, fractal speckles are also analyzed to examine the self-similar properties of the resulting field. The correlation properties of clipped speckles are investigated to explore the possibility of fabricating fractal random media by means of photopolymerization for three-dimensional patterning of micro structures.

  6. Experiment research on orthogonal tilting scanner

    NASA Astrophysics Data System (ADS)

    Li, Anhu; Liu, Liren; Sun, Jianfeng; Zhong, Xianghong; Luan, Zhu

    2007-09-01

    The original scanner of tilting orthogonal double prisms is studied for testing the tracking performance in inter-satellite laser communications. Two prisms respectively rotate around the horizontal axle and the vertical one within the admissible range to determine the corresponding orientation and position of the passing beam, therefore the high accuracy deviation angle of passing beam can be performed. The test experiments performed with autocollimator and interferometer, as well as the theoretical analysis, indicates that the scanner can meet the requirements of the deviation accuracy superior to 0.5 μrad with the deviation range greater than 500 μrad, which accords to our design requirements.

  7. Noncommutative Pfaffians associated with the orthogonal algebra

    SciTech Connect

    Artamonov, Dmitrii V; Golubeva, Valentina A

    2012-12-31

    Commutators of Pfaffians associated with the orthogonal algebra are found in skew-symmetric and root realizations of o{sub N}. A generating function of Pfaffians is proved to satisfy the reflection equation. A relation between Pfaffians in skew-symmetric and root realizations of o{sub N} is established. Using these results we construct an integrable equation of Knizhnik-Zamolodchikov type using the Capelli central elements in U(o{sub N}), which are sums of squares of the considered Pfaffians. A classical limit of the obtained Knizhnik-Zamolodchikov type equation turns out to be a very specific system of equations of isomonodromic deformations. Bibliography: 18 titles.

  8. Functional expansion of human tRNA synthetases achieved by structural inventions.

    PubMed

    Guo, Min; Schimmel, Paul; Yang, Xiang-Lei

    2010-01-21

    Known as an essential component of the translational apparatus, the aminoacyl-tRNA synthetase family catalyzes the first step reaction in protein synthesis, that is, to specifically attach each amino acid to its cognate tRNA. While preserving this essential role, tRNA synthetases developed other roles during evolution. Human tRNA synthetases, in particular, have diverse functions in different pathways involving angiogenesis, inflammation and apoptosis. The functional diversity is further illustrated in the association with various diseases through genetic mutations that do not affect aminoacylation or protein synthesis. Here we review the accumulated knowledge on how human tRNA synthetases used structural inventions to achieve functional expansions.

  9. Orthogonally-oriented nanotube arrays: experiment I.

    PubMed

    Sheehan, D P; Webster, J T; Baird, L M

    2007-10-01

    Recently a new type of self-assembling surface has been proposed that, in theory, possesses a number of desirable tribological, electrical, and thermal characterstics. The surface consists of arrays of carbon nanotubes partially embedded lengthwise in a substrate such that when two arrayed surfaces are brought together orthogonally, the areal contact between them is small, limited to a lattice of nearly point-like contacts. These orthogonally-oriented nanotube arrays (ONAs) are predicted to exhibit: (i) surface adhesion (stiction) 10-100 times less than for Teflon or other advanced perfluorocarbons; (ii) frictional coefficients up to 1000 times less than for conventional solids; (iii) ultra-low wear; and (iv) superior thermal and electrical conductivity. In this paper, laboratory methods are described for embedding nanotubes in trenched substrates. Using microscopically trenched substrates and a custom ultrasonic atomization source, experiments show that individual nanotubes can spontaneously and controllably entrench themselves via interfacial forces (capillary and surface tension). Results indicate ONAs might be relatively simply and inexpensively fabricated. More decisive experiments are proposed.

  10. Regulation of glutamine synthetase, aspartokinase, and total protein turnover in Klebsiella aerogenes.

    PubMed

    Fulks, R M; Stadtman, E R

    1985-12-13

    When suspensions of Klebsiella aerogenes are incubated in a nitrogen-free medium there is a gradual decrease in the levels of acid-precipitable protein and of aspartokinase III (lysine-sensitive) and aspartokinase I (threonine-sensitive) activities. In contrast, the level of glutamine synthetase increases slightly and then remains constant. Under these conditions, the glutamine synthetase and other proteins continue to be synthesized as judged by the incorporation of [14C]leucine into the acid-precipitable protein fraction and into protein precipitated by anti-glutamine synthetase antibodies, by the fact that growth-inhibiting concentrations of chloramphenicol also inhibit the incorporation of [14C]leucine into protein and into protein precipitated by anti-glutamine synthetase antibody, and by the fact that chloramphenicol leads to acceleration in the loss of aspartokinases I and III and promotes a net decrease in the level of glutamine synthetase and its cross-reactive protein. The loss of aspartokinases I and III in cell suspensions is stimulated by glucose and is inhibited by 2,4-dinitrophenol. Glucose also stimulates the loss of aspartokinases and glutamine synthetase in the presence of chloramphenicol. Cell-free extracts of K. aerogenes catalyze rapid inactivation of endogenous glutamine synthetase as well as exogenously added pure glutamine synthetase. This loss of glutamine synthetase is not associated with a loss of protein that cross-reacts with anti-glutamine synthetase antibodies. The inactivation of glutamine synthetase in extracts is not due to adenylylation. It is partially prevented by sulfhydryl reagents, Mn2+, antimycin A, 2,4-dinitrophenol, EDTA, anaerobiosis and by dialysis. Following 18 h dialysis, the capacity of extracts to catalyze inactivation of glutamine synthetase is lost but can be restored by the addition of Fe2+ (or Ni2+) together with ATP (or other nucleoside di- and triphosphates. After 40-60 h dialysis Fe3+ together with NADH (but

  11. Assignment of the cysteinyl-tRNA synthetase gene (CARS) to 11p15. 5

    SciTech Connect

    Cruzen, M.E.; Bengtsson, U.; McMahon, J.; Wasmuth, J.J.; Arfin, S.M. )

    1993-03-01

    The attachment of each of the 20 naturally occurring amino acids to their cognate tRNA isoaccepting families is catalyzed by a specific aminoacyl-tRNA synthetase. The structural genes encoding 10 of these enzymes have been assigned to specific human chromosomes. The HARS, LARS, RARS, and TARS genes, encoding histidyl-, leucyl-, arginyl-, and threonyl-tRNA synthetases, respectively, are all located on chromosome 5( 1, 5, 7, 9, 14). The MARS (methionyl-tRNA synthetase), NARS (asparaginyl-tRNA synthetase), VARS (valyl-tRNA synthetase), and WARS (tryptophanyl-tRNA synthetase) genes have been assigned to chromosomes 12, 18, 6, and 14, respectively (3, 4, 6, 8). A gene originally identified as encoding glutaminyl-tRNA synthetase was mapped to chromosome 1q32-q42 (10). However, a recent study suggests that the product of this gene is, in fact, a multifunctional enzyme with both glutamyl- and prolyl-tRNA synthetase activities (2). The fact that 4 of the 10 aminoacyl-tRNA synthetase genes already mapped are located on chromosome 5 may be fortuitous but might also indicate an evolutionary or regulatory relatedness. It is therefore, of interest to map genes encoding other aminoacyl-tRNA synthetases to determine if additional examples of synteny exist. The recent isolation of cDNA and genomic DNA clones for human cysteinyl-tRNA synthetase has now enabled us to map the CARS gene to segment p15.5 on chromosome 11 by fluorescence in situ hybridization.

  12. Taguchi’s Orthogonal Arrays Are Classical Designs of Experiments

    PubMed Central

    Kacker, Raghu N.; Lagergren, Eric S.; Filliben, James J.

    1991-01-01

    Taguchi’s catalog of orthogonal arrays is based on the mathematical theory of factorial designs and difference sets developed by R. C. Bose and his associates. These arrays evolved as extensions of factorial designs and latin squares. This paper (1) describes the structure and constructions of Taguchi’s orthogonal arrays, (2) illustrates their fractional factorial nature, and (3) points out that Taguchi’s catalog can be expanded to include orthogonal arrays developed since 1960. PMID:28184132

  13. Orthogonal fast spherical Bessel transform on uniform grid

    NASA Astrophysics Data System (ADS)

    Serov, Vladislav V.

    2017-07-01

    We propose an algorithm for the orthogonal fast discrete spherical Bessel transform on a uniform grid. Our approach is based upon the spherical Bessel transform factorization into the two subsequent orthogonal transforms, namely the fast Fourier transform and the orthogonal transform founded on the derivatives of the discrete Legendre orthogonal polynomials. The method utility is illustrated by its implementation for the problem of a two-atomic molecule in a time-dependent external field simulating the one utilized in the attosecond streaking technique.

  14. Orthogonal patterns in binary neural networks

    NASA Technical Reports Server (NTRS)

    Baram, Yoram

    1988-01-01

    A binary neural network that stores only mutually orthogonal patterns is shown to converge, when probed by any pattern, to a pattern in the memory space, i.e., the space spanned by the stored patterns. The latter are shown to be the only members of the memory space under a certain coding condition, which allows maximum storage of M=(2N) sup 0.5 patterns, where N is the number of neurons. The stored patterns are shown to have basins of attraction of radius N/(2M), within which errors are corrected with probability 1 in a single update cycle. When the probe falls outside these regions, the error correction capability can still be increased to 1 by repeatedly running the network with the same probe.

  15. Partially orthogonal resonators for magnetic resonance imaging

    NASA Astrophysics Data System (ADS)

    Chacon-Caldera, Jorge; Malzacher, Matthias; Schad, Lothar R.

    2017-02-01

    Resonators for signal reception in magnetic resonance are traditionally planar to restrict coil material and avoid coil losses. Here, we present a novel concept to model resonators partially in a plane with maximum sensitivity to the magnetic resonance signal and partially in an orthogonal plane with reduced signal sensitivity. Thus, properties of individual elements in coil arrays can be modified to optimize physical planar space and increase the sensitivity of the overall array. A particular case of the concept is implemented to decrease H-field destructive interferences in planar concentric in-phase arrays. An increase in signal to noise ratio of approximately 20% was achieved with two resonators placed over approximately the same planar area compared to common approaches at a target depth of 10 cm at 3 Tesla. Improved parallel imaging performance of this configuration is also demonstrated. The concept can be further used to increase coil density.

  16. Measuring the orthogonality error of coil systems

    USGS Publications Warehouse

    Heilig, B.; Csontos, A.; Pajunpää, K.; White, Tim; St. Louis, B.; Calp, D.

    2012-01-01

    Recently, a simple method was proposed for the determination of pitch angle between two coil axes by means of a total field magnetometer. The method is applicable when the homogeneous volume in the centre of the coil system is large enough to accommodate the total field sensor. Orthogonality of calibration coil systems used for calibrating vector magnetometers can be attained by this procedure. In addition, the method can be easily automated and applied to the calibration of delta inclination–delta declination (dIdD) magnetometers. The method was tested by several independent research groups, having a variety of test equipment, and located at differing geomagnetic observatories, including: Nurmijärvi, Finland; Hermanus, South Africa; Ottawa, Canada; Tihany, Hungary. This paper summarizes the test results, and discusses the advantages and limitations of the method.

  17. Response Surface Modeling Using Multivariate Orthogonal Functions

    NASA Technical Reports Server (NTRS)

    Morelli, Eugene A.; DeLoach, Richard

    2001-01-01

    A nonlinear modeling technique was used to characterize response surfaces for non-dimensional longitudinal aerodynamic force and moment coefficients, based on wind tunnel data from a commercial jet transport model. Data were collected using two experimental procedures - one based on modem design of experiments (MDOE), and one using a classical one factor at a time (OFAT) approach. The nonlinear modeling technique used multivariate orthogonal functions generated from the independent variable data as modeling functions in a least squares context to characterize the response surfaces. Model terms were selected automatically using a prediction error metric. Prediction error bounds computed from the modeling data alone were found to be- a good measure of actual prediction error for prediction points within the inference space. Root-mean-square model fit error and prediction error were less than 4 percent of the mean response value in all cases. Efficacy and prediction performance of the response surface models identified from both MDOE and OFAT experiments were investigated.

  18. Inverse solutions for tilting orthogonal double prisms.

    PubMed

    Li, Anhu; Ding, Ye; Bian, Yongming; Liu, Liren

    2014-06-10

    An analytical reverse solution and actual examples are given to show how to direct a laser beam from a pair of orthogonal prisms to given targets in free space. Considering the influences of double-prism structural parameters, a lookup table method to seek the numerical reverse solution of each prism's tilting angle is also proposed for steering the double-prism orientation to track a target position located in the near field. Some case studies, as well as a specified elliptical target trajectory scanned by the cam-based driving double prisms, exhibit the significant application values of the theoretical derivation. The analytic reverse and numerical solutions can be generalized to investigate the synthesis of scanning patterns and the controlling strategy of double-prism tilting motion, the potentials of which can be explored to perform the orientation and position tracking functions in applications of precision engineering fields.

  19. Partially orthogonal resonators for magnetic resonance imaging

    PubMed Central

    Chacon-Caldera, Jorge; Malzacher, Matthias; Schad, Lothar R.

    2017-01-01

    Resonators for signal reception in magnetic resonance are traditionally planar to restrict coil material and avoid coil losses. Here, we present a novel concept to model resonators partially in a plane with maximum sensitivity to the magnetic resonance signal and partially in an orthogonal plane with reduced signal sensitivity. Thus, properties of individual elements in coil arrays can be modified to optimize physical planar space and increase the sensitivity of the overall array. A particular case of the concept is implemented to decrease H-field destructive interferences in planar concentric in-phase arrays. An increase in signal to noise ratio of approximately 20% was achieved with two resonators placed over approximately the same planar area compared to common approaches at a target depth of 10 cm at 3 Tesla. Improved parallel imaging performance of this configuration is also demonstrated. The concept can be further used to increase coil density. PMID:28186135

  20. Partially orthogonal resonators for magnetic resonance imaging.

    PubMed

    Chacon-Caldera, Jorge; Malzacher, Matthias; Schad, Lothar R

    2017-02-10

    Resonators for signal reception in magnetic resonance are traditionally planar to restrict coil material and avoid coil losses. Here, we present a novel concept to model resonators partially in a plane with maximum sensitivity to the magnetic resonance signal and partially in an orthogonal plane with reduced signal sensitivity. Thus, properties of individual elements in coil arrays can be modified to optimize physical planar space and increase the sensitivity of the overall array. A particular case of the concept is implemented to decrease H-field destructive interferences in planar concentric in-phase arrays. An increase in signal to noise ratio of approximately 20% was achieved with two resonators placed over approximately the same planar area compared to common approaches at a target depth of 10 cm at 3 Tesla. Improved parallel imaging performance of this configuration is also demonstrated. The concept can be further used to increase coil density.

  1. Description of sunspot cycles by orthogonal functions

    NASA Technical Reports Server (NTRS)

    Teuber, D. L.; Reichmann, E. J.; Wilson, R. M.

    1984-01-01

    Based on the principal component analysis technique and evidence for a 22-yr double-sunspot cycle periodicity. The time series of sunspot numbers is represented as a sum of mutually orthogonal eigenvectors in the time domain. It is shown that the first two eigenvectors account for about 90 percent of the cumulative 'signal power,' and that this is sufficient for reconstruction of the raw data curve. It is also noted that the second eigenvector behaves as the time derivative of the first, and that a phase-plane plot of these eigenvectors (i.e. a plot of a variable vs. its rate of change) suggests that the sun's sunspot cycle is driven by an oscillator; the implication is that, embedded within the sun, a chronometer is at work (e.g. Dicke, 1979).

  2. Nanomachining of non-orthogonal mask patterns

    NASA Astrophysics Data System (ADS)

    Robinson, Tod; Yi, Daniel; White, Roy; Bozak, Ron; Archuletta, Mike; Lee, David

    2010-05-01

    Patterns which are not aligned to standard orthogonal (x and y ordinate) directions have recently been developed for advanced lithography nodes. Efforts have been successful in developing single pass nanomachining repair processes to meet the printability requirements for these patterns. This development makes use of the latest improvements made to the COBRA repair process (the Enhanced COBRA process typically completed in less than 2 minutes of repair time) with symmetric NanoBits to repair opposing critical edges of bridging defects. It also required fundamental changes in the software tools to allow automated detection of the angle of the edges and the application of pre-programmed repair edge biases normal (90°) to the detected angled edges. Additionally, some other new improvements (hardware, software, and process) are reviewed in light of more traditional nanomachining repairs.

  3. Inactivation and covalent modification of CTP synthetase by thiourea dioxide.

    PubMed Central

    Robertson, J. G.; Sparvero, L. J.; Villafranca, J. J.

    1992-01-01

    Thiourea dioxide was used in chemical modification studies to identify functionally important amino acids in Escherichia coli CTP synthetase. Incubation at pH 8.0 in the absence of substrates led to rapid, time dependent, and irreversible inactivation of the enzyme. The second-order rate constant for inactivation was 0.18 M-1 s-1. Inactivation also occurred in the absence of oxygen and in the presence of catalase, thereby ruling out mixed-function oxidation/reduction as the mode of amino acid modification. Saturating concentrations of the substrates ATP and UTP, and the allosteric activator GTP prevented inactivation by thiourea dioxide, whereas saturating concentrations of glutamine (a substrate) did not. The concentration dependence of nucleotide protection revealed cooperative behavior with respect to individual nucleotides and with respect to various combinations of nucleotides. Mixtures of nucleotides afforded greater protection against inactivation than single nucleotides alone, and a combination of the substrates ATP and UTP provided the most protection. The Hill coefficient for nucleotide protection was approximately 2 for ATP, UTP, and GTP. In the presence of 1:1 ratios of ATP:UTP, ATP:GTP, and UTP:GTP, the Hill coefficient was approximately 4 in each case. Fluorescence and circular dichroism measurements indicated that modification by thiourea dioxide causes detectable changes in the structure of the protein. Modification with [14C]thiourea dioxide demonstrated that complete inactivation correlates with incorporation of 3 mol of [14C]thiourea dioxide per mole of CTP synthetase monomer. The specificity of thiourea dioxide for lysine residues indicates that one or more lysines are most likely involved in CTP synthetase activity. The data further indicate that nucleotide binding prevents access to these functionally important residues. PMID:1303749

  4. Mammalian long-chain acyl-CoA synthetases.

    PubMed

    Soupene, Eric; Kuypers, Frans A

    2008-05-01

    Acyl-CoA synthetase enzymes are essential for de novo lipid synthesis, fatty acid catabolism, and remodeling of membranes. Activation of fatty acids requires a two-step reaction catalyzed by these enzymes. In the first step, an acyl-AMP intermediate is formed from ATP. AMP is then exchanged with CoA to produce the activated acyl-CoA. The release of AMP in this reaction defines the superfamily of AMP-forming enzymes. The length of the carbon chain of the fatty acid species defines the substrate specificity for the different acyl-CoA synthetases (ACS). On this basis, five sub-families of ACS have been characterized. The purpose of this review is to report on the large family of mammalian long-chain acyl-CoA synthetases (ACSL), which activate fatty acids with chain lengths of 12 to 20 carbon atoms. Five genes and several isoforms generated by alternative splicing have been identified and limited information is available on their localization. The structure of these membrane proteins has not been solved for the mammalian ACSLs but homology to a bacterial form, whose structure has been determined, points at specific structural features that are important for these enzymes across species. The bacterial form acts as a dimer and has a conserved short motif, called the fatty acid Gate domain, that seems to determine substrate specificity. We will discuss the characterization and identification of the different spliced isoforms, draw attention to the inconsistencies and errors in their annotations, and their cellular localizations. These membrane proteins act on membrane-bound substrates probably as homo- and as heterodimer complexes but have often been expressed as single recombinant isoforms, apparently purified as monomers and tested in Triton X-100 micelles. We will argue that such studies have failed to provide an accurate assessment of the activity and of the distinct function of these enzymes in mammalian cells.

  5. Phosphinothricin Tripeptide Synthetases in Streptomyces viridochromogenes Tü494

    PubMed Central

    Schwartz, Dirk; Grammel, Nicolas; Heinzelmann, Eva; Keller, Ullrich; Wohlleben, Wolfgang

    2005-01-01

    The tripeptide backbone of phosphinothricin (PT) tripeptide (PTT), a compound with herbicidal activity from Streptomyces viridochromogenes, is assembled by three stand-alone peptide synthetase modules. The enzyme PhsA (66 kDa) recruits the PT-precursor N-acetyl-demethylphosphinothricin (N-Ac-DMPT), whereas the two alanine residues of PTT are assembled by the enzymes PhsB and PhsC (129 and 119 kDa, respectively). During or after assembly, the N-Ac-DMPT residue in the peptide is converted to PT by methylation and deacetylation. Both phsB and phsC appear to be cotranscribed together with two other genes from a single promoter and they are located at a distance of 20 kb from the gene phsA, encoding PhsA, in the PTT biosynthesis gene cluster of S. viridochromogenes. PhsB and PhsC represent single nonribosomal peptide synthetase elongation modules lacking a thioesterase domain. Gene inactivations, genetic complementations, determinations of substrate specificity of the heterologously produced proteins, and comparison of PhsC sequence with the amino terminus of the alanine-activating nonribosomal peptide synthetase PTTSII from S. viridochromogenes confirmed the role of the two genes in the bialanylation of Ac-DMPT. The lack of an integral thioesterase domain in the PTT assembly system points to product release possibly involving two type II thioesterase genes (the1 and the2) located in the PTT gene cluster alone or in conjunction with an as yet unknown mechanism of product release. PMID:16251301

  6. Glutamine versus Ammonia Utilization in the NAD Synthetase Family

    PubMed Central

    Shatalin, Konstantin; Gelfand, Mikhail S.; Osterman, Andrei L.; Sorci, Leonardo

    2012-01-01

    NAD is a ubiquitous and essential metabolic redox cofactor which also functions as a substrate in certain regulatory pathways. The last step of NAD synthesis is the ATP-dependent amidation of deamido-NAD by NAD synthetase (NADS). Members of the NADS family are present in nearly all species across the three kingdoms of Life. In eukaryotic NADS, the core synthetase domain is fused with a nitrilase-like glutaminase domain supplying ammonia for the reaction. This two-domain NADS arrangement enabling the utilization of glutamine as nitrogen donor is also present in various bacterial lineages. However, many other bacterial members of NADS family do not contain a glutaminase domain, and they can utilize only ammonia (but not glutamine) in vitro. A single-domain NADS is also characteristic for nearly all Archaea, and its dependence on ammonia was demonstrated here for the representative enzyme from Methanocaldococcus jannaschi. However, a question about the actual in vivo nitrogen donor for single-domain members of the NADS family remained open: Is it glutamine hydrolyzed by a committed (but yet unknown) glutaminase subunit, as in most ATP-dependent amidotransferases, or free ammonia as in glutamine synthetase? Here we addressed this dilemma by combining evolutionary analysis of the NADS family with experimental characterization of two representative bacterial systems: a two-subunit NADS from Thermus thermophilus and a single-domain NADS from Salmonella typhimurium providing evidence that ammonia (and not glutamine) is the physiological substrate of a typical single-domain NADS. The latter represents the most likely ancestral form of NADS. The ability to utilize glutamine appears to have evolved via recruitment of a glutaminase subunit followed by domain fusion in an early branch of Bacteria. Further evolution of the NADS family included lineage-specific loss of one of the two alternative forms and horizontal gene transfer events. Lastly, we identified NADS structural

  7. Structural analysis of FAD synthetase from Corynebacterium ammoniagenes

    PubMed Central

    Frago, Susana; Martínez-Júlvez, Marta; Serrano, Ana; Medina, Milagros

    2008-01-01

    Background The prokaryotic FAD synthetase family – a group of bifunctional enzymes that catalyse riboflavin phosphorylation and FMN adenylylation within a single polypeptide chain- was analysed in terms of sequence and structure. Results Sequences of nearly 800 prokaryotic species were aligned. Those related with bifunctional FAD synthetase activities showed conservation of several consensus regions and highly conserved residues. A 3D model for the FAD synthetase from Corynebacterium ammoniagenes (CaFADS) was generated. This model confirms that the N-terminal and C-terminal domains are related to nucleotydyltransferases and riboflavin kinases, respectively. Models for the interaction of CaFADS with its substrates were also produced, allowing location of all the protein substrates in their putative binding pockets. These include two independent flavin binding sites for each CaFADS activity. Conclusion For the first time, the putative presence of a flavin binding site for the adenylylation activity, independent from that related with the phosphorylation activity, is shown. Additionally, these models suggest the functional relevance of some residues putatively involved in the catalytic processes. Their relevant roles were analysed by site-directed mutagenesis. A role was confirmed for H28, H31, S164 and T165 in the stabilisation of the P groups and the adenine moiety of ATP and, the P of FMN for the adenylylation. Similarly, T208, N210 and E268 appear critical for accommodation of the P groups of ATP and the ribityl end of RF in the active site for the phosphorylation process. Finally, the C-terminal domain was shown to catalyse the phosphorylation process on its own, but no reaction at all was observed with the individually expressed N-terminal domain. PMID:18811972

  8. Structural analysis of FAD synthetase from Corynebacterium ammoniagenes.

    PubMed

    Frago, Susana; Martínez-Júlvez, Marta; Serrano, Ana; Medina, Milagros

    2008-09-23

    The prokaryotic FAD synthetase family - a group of bifunctional enzymes that catalyse riboflavin phosphorylation and FMN adenylylation within a single polypeptide chain- was analysed in terms of sequence and structure. Sequences of nearly 800 prokaryotic species were aligned. Those related with bifunctional FAD synthetase activities showed conservation of several consensus regions and highly conserved residues. A 3D model for the FAD synthetase from Corynebacterium ammoniagenes (CaFADS) was generated. This model confirms that the N-terminal and C-terminal domains are related to nucleotydyltransferases and riboflavin kinases, respectively. Models for the interaction of CaFADS with its substrates were also produced, allowing location of all the protein substrates in their putative binding pockets. These include two independent flavin binding sites for each CaFADS activity. For the first time, the putative presence of a flavin binding site for the adenylylation activity, independent from that related with the phosphorylation activity, is shown. Additionally, these models suggest the functional relevance of some residues putatively involved in the catalytic processes. Their relevant roles were analysed by site-directed mutagenesis. A role was confirmed for H28, H31, S164 and T165 in the stabilisation of the P groups and the adenine moiety of ATP and, the P of FMN for the adenylylation. Similarly, T208, N210 and E268 appear critical for accommodation of the P groups of ATP and the ribityl end of RF in the active site for the phosphorylation process. Finally, the C-terminal domain was shown to catalyse the phosphorylation process on its own, but no reaction at all was observed with the individually expressed N-terminal domain.

  9. LOCC indistinguishable orthogonal product quantum states

    PubMed Central

    Zhang, Xiaoqian; Tan, Xiaoqing; Weng, Jian; Li, Yongjun

    2016-01-01

    We construct two families of orthogonal product quantum states that cannot be exactly distinguished by local operation and classical communication (LOCC) in the quantum system of 2k+i ⊗ 2l+j (i, j ∈ {0, 1} and i ≥ j ) and 3k+i ⊗ 3l+j (i, j ∈ {0, 1, 2}). And we also give the tiling structure of these two families of quantum product states where the quantum states are unextendible in the first family but are extendible in the second family. Our construction in the quantum system of 3k+i ⊗ 3l+j is more generalized than the other construction such as Wang et al.’s construction and Zhang et al.’s construction, because it contains the quantum system of not only 2k ⊗ 2l and 2k+1 ⊗ 2l but also 2k ⊗ 2l+1 and 2k+1 ⊗ 2l+1. We calculate the non-commutativity to quantify the quantumness of a quantum ensemble for judging the local indistinguishability. We give a general method to judge the indistinguishability of orthogonal product states for our two constructions in this paper. We also extend the dimension of the quantum system of 2k ⊗ 2l in Wang et al.’s paper. Our work is a necessary complement to understand the phenomenon of quantum nonlocality without entanglement. PMID:27377310

  10. LOCC indistinguishable orthogonal product quantum states

    NASA Astrophysics Data System (ADS)

    Zhang, Xiaoqian; Tan, Xiaoqing; Weng, Jian; Li, Yongjun

    2016-07-01

    We construct two families of orthogonal product quantum states that cannot be exactly distinguished by local operation and classical communication (LOCC) in the quantum system of 2k+i ⊗ 2l+j (i, j ∈ {0, 1} and i ≥ j ) and 3k+i ⊗ 3l+j (i, j ∈ {0, 1, 2}). And we also give the tiling structure of these two families of quantum product states where the quantum states are unextendible in the first family but are extendible in the second family. Our construction in the quantum system of 3k+i ⊗ 3l+j is more generalized than the other construction such as Wang et al.’s construction and Zhang et al.’s construction, because it contains the quantum system of not only 2k ⊗ 2l and 2k+1 ⊗ 2l but also 2k ⊗ 2l+1 and 2k+1 ⊗ 2l+1. We calculate the non-commutativity to quantify the quantumness of a quantum ensemble for judging the local indistinguishability. We give a general method to judge the indistinguishability of orthogonal product states for our two constructions in this paper. We also extend the dimension of the quantum system of 2k ⊗ 2l in Wang et al.’s paper. Our work is a necessary complement to understand the phenomenon of quantum nonlocality without entanglement.

  11. Stability of Rat Brain Glutamine Synthetase to Oxygen Toxicity (Oxygen at High Pressure).

    DTIC Science & Technology

    1983-07-01

    Enzyme assays using the gamma-glutamyl transferase method provided estimates of glutamine synthetase activity in rat brain homogenates subjected to a...supports the lack of any connection between convulsions caused by in vivo inhibition of glutamine synthetase and convulsions caused by oxygen toxicity (oxygen at high pressure). (Author)

  12. Peptide Mapping of Aminoacyl-tRNA Synthetases: Evidence for Internal Sequence Homology in Escherichia coli Leucyl-tRNA Synthetase

    PubMed Central

    Waterson, Robert M.; Konigsberg, William H.

    1974-01-01

    Most aminoacyl-tRNA synthetases contain polypeptide chains of about either 50,000 or 100,000 daltons. Peptide mapping of tryptic, chymotryptic, or Staphylococcus aureus acid protease digests of seryl-tRNA synthetase (100,000, dimer) and leucyl-tRNA synthetase (100,000, monomer) from E. coli was done after selective modification of lysine residues with [14C]succinic anhydride or of methionine residues with [14C]iodoacetate. By use of thin-layer electrophoresis and chromatography on silicagel or cellulose plates followed by radioautography it was possible, depending upon the specific activity of the reagent used, to detect radioactive peptides obtained from as little as l μg of protein. Seryl-tRNA synthetase gave the correct number of tryptic peptides expected for a dimer of identical subunits. Leucyl-tRNA synthetase, on the other hand, gave roughly half the number of radioactive tryptic, chymotryptic, and acid protease peptides expected from the lysine, arginine, and methionine content of the 100,000 monomer. We have interpreted these results as indicating that extensive internal homology exists among lysine- and methionine-containing peptides within the leucyl-tRNA synthetase. The simplest conclusion that can be drawn from these observations is that the NH2- and COOH-terminal halves of leucyl-tRNA synthetase and perhaps other synthetases of 100,000 molecular weight may have evolved through a process of gene duplication and fusion, followed by limited diversification by way of amino-acid substitutions accumulating during evolution. Images PMID:4592690

  13. Circumstantial evidence for a role of glutamine-synthetase in suicide.

    PubMed

    Kalkman, Hans O

    2011-06-01

    Suicide occurs during depression, schizophrenia, diabetes and epilepsy. A common denominator of these disorders is the presence of inflammation. Inflammatory cytokines affect function and expression of the glial enzyme glutamine synthetase and post mortem studies indicate that brain glutamine synthetase function is suppressed in mood disorders and epilepsy. In a study of schizophrenia brains, the expression of glutamine synthetase was reduced in those cases where the cause of death was suicide. The glycogen synthase kinase 3 (GSK3) inhibitor, lithium, which has a proven efficacy against suicide, increased in an animal experiment the expression of glutamine synthetase. Based on these data one could reason that suicide may be prevented by centrally acting GSK3 inhibitors. However, since inhibition of glutamine synthetase may lead to a deficit in glutamine and as consequence a GABA and glutamate deficit, even simple food supplementation with glutamine might help to reduce suicide.

  14. Myocardial aminoacyl-transfer-ribonucleic acid synthetase and aminoacyl-transferring enzyme activity

    PubMed Central

    Gibson, K.; Harris, P.

    1972-01-01

    The properties of cytoplasmic aminoacyl-tRNA synthetase and aminoacyl-transferring enzymes in the myocardium were examined and methods for the assay of the activity of these enzyme systems were developed. Aminoacyl-tRNA synthetase activity was measured from the rate of incorporation of 14C-labelled amino acid into aminoacyl-tRNA. Transferase activity was measured from the rate of incorporation of amino[14C]acyl-tRNA into protein in the presence of a standard preparation of hepatic ribosomes. Aminoacyl-tRNA synthetase activity is labile once the heart has been homogenized, whereas transferase activity is stable. The source of energy for synthetase activity is ATP; that for transferase is GTP. Transferase activity was inhibited by puromycin and stimulated by dithiothreitol, whereas synthetase activity was unaffected. PMID:5071178

  15. Inhibition of rabbit gastric glucosamine synthetase activity by Cu2+, Zn2+ and Se4+.

    PubMed

    Fujita, T; Sakuma, S; Takahashi, K; Bohtani, Y; Nishida, H; Fujimoto, Y

    1997-05-01

    The effects of Fe2+, Cu2+, Zn2+ and Se4+ on the activity of glucosamine synthetase, the rate-limiting enzyme of mucus synthesis, in rabbit gastric corporal mucosa were examined. Cu2+, Zn2+ and Se4+ inhibited the glucosamine synthetase activity at concentrations ranging from 1 to 10 microM (Cu2+, 8-98% inhibition; Zn2+, 10-99% inhibition; Se4+, 32-89% inhibition). The inhibitory effects of these three ions were much stronger than that of UDP-N-acetylglúcosamine known as a representative inhibitor of the glucosamine synthetase activity (10 microM, 52% inhibition). Fe2+ had no significant effect on the glucosamine synthetase activity up to 100 microM. These results suggest that Cu2+, Zn2+ and Se4+ can be potent inhibitors of gastric glucosamine synthetase activity.

  16. The evolution of Class II Aminoacyl-tRNA synthetases and the first code.

    PubMed

    Smith, Temple F; Hartman, Hyman

    2015-11-30

    Class II Aminoacyl-tRNA synthetases are a set of very ancient multi domain proteins. The evolution of the catalytic domain of Class II synthetases can be reconstructed from three peptidyl-hairpins. Further evolution from this primordial catalytic core leads to a split of the Class II synthetases into two divisions potentially associated with the operational code. The earliest form of this code likely coded predominantly Glycine (Gly), Proline (Pro), Alanine (Ala) and "Lysine"/Aspartic acid (Lys/Asp). There is a paradox in these synthetases beginning with a hairpin structure before the Genetic Code existed. A resolution is found in the suggestion that the primordial Aminoacyl synthetases formed in a transition from a Thioester world to a Phosphate ester world. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  17. Replacement of the folC gene, encoding folylpolyglutamate synthetase-dihydrofolate synthetase in Escherichia coli, with genes mutagenized in vitro.

    PubMed Central

    Pyne, C; Bognar, A L

    1992-01-01

    The folylpolyglutamate synthetase-dihydrofolate synthetase gene (folC) in Escherichia coli was deleted from the bacterial chromosome and replaced by a selectable Kmr marker. The deletion strain required a complementing gene expressing folylpolyglutamate synthetase encoded on a plasmid for viability, indicating that folC is an essential gene in E. coli. The complementing folC gene was cloned into the vector pPM103 (pSC101, temperature sensitive for replication), which segregated spontaneously at 42 degrees C in the absence of selection. This complementing plasmid was replaced in the folC deletion strain by compatible pUC plasmids containing folC genes with mutations generated in vitro, producing strains which express only mutant folylpolyglutamate synthetase. Mutant folC genes expressing insufficient enzyme activity could not complement the chromosomal deletion, resulting in retention of the pPM103 plasmid. Some mutant genes expressing low levels of enzyme activity replaced the complementing plasmid, but the strains produced were auxotrophic for products of folate-dependent pathways. The folylpolyglutamate synthetase gene from Lactobacillus casei, which may lack dihydrofolate synthetase activity, replaced the complementing plasmid, but the strain was auxotrophic for all folate end products. Images PMID:1548226

  18. Purification and comparison of two forms of S-adenosyl-L-methionine synthetase from rat liver.

    PubMed

    Cabrero, C; Puerta, J; Alemany, S

    1987-12-30

    Only two S-adenosyl-L-methionine synthetase forms exist in rat liver: high-Mr S-adenosyl-L-methionine synthetase and low-Mr S-adenosyl-L-methionine synthetase, which have been purified to apparent homogeneity as judged by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. High-Mr S-adenosyl-L-methionine synthetase had an apparent molecular mass, determined by gel filtration, of 210 kDa and was a tetramer constituted by 48.5-kDa subunits, estimated by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The apparent molecular mass of low-Mr S-adenosyl-L-methionine synthetase, as estimated by gel filtration, was 110 kDa and was constituted by two subunits of 47 kDa. An antiserum against low-Mr S-adenosyl-L-methionine synthetase cross-reacted with the two forms. Reverse-phase HPLC runs of tryptic digestions of high-Mr and low-Mr S-adenosyl-L-methionine synthetase showed that the peptide maps of the two forms were very similar, if not identical. High-Mr S-adenosyl-L-methionine synthetase activity was inhibited by S-adenosyl-L-methionine and pyrophosphate. Depending on the dose used, S-adenosyl-L-methionine activated or inhibited low-Mr S-adenosyl-L-methionine synthetase and pyrophosphate had no effect on this form. The two synthetases showed a different specific activity at the physiological concentration of methionine. This report shows that even though the two forms are constructed of the same polypeptide chains, they are regulated in a different manner by methionine and by the products of the reaction.

  19. Novel insights into regulation of asparagine synthetase in conifers.

    PubMed

    Canales, Javier; Rueda-López, Marina; Craven-Bartle, Blanca; Avila, Concepción; Cánovas, Francisco M

    2012-01-01

    Asparagine, a key amino acid for nitrogen storage and transport in plants, is synthesized via the ATP-dependent reaction catalyzed by the enzyme asparagine synthetase (AS; EC 6.3.5.4). In this work, we present the molecular analysis of two full-length cDNAs that encode asparagine synthetase in maritime pine (Pinus pinaster Ait.), PpAS1, and PpAS2. Phylogenetic analyses of the deduced amino acid sequences revealed that both genes are class II AS, suggesting an ancient origin of these genes in plants. A comparative study of PpAS1 and PpAS2 gene expression profiles showed that PpAS1 gene is highly regulated by developmental and environmental factors, while PpAS2 is expressed constitutively. To determine the molecular mechanisms underpinning the differential expression of PpAS1, the promoter region of the gene was isolated and putative binding sites for MYB transcription factors were identified. Gel mobility shift assays showed that a MYB protein from Pinus taeda (PtMYB1) was able to interact with the promoter region of PpAS1. Furthermore, transient expression analyses in pine cells revealed a negative effect of PtMYB1 on PpAS1 expression. The potential role of MYB factors in the transcriptional regulation of PpAS1 in vascular cells is discussed.

  20. Novel Insights into Regulation of Asparagine Synthetase in Conifers

    PubMed Central

    Canales, Javier; Rueda-López, Marina; Craven-Bartle, Blanca; Avila, Concepción; Cánovas, Francisco M.

    2012-01-01

    Asparagine, a key amino acid for nitrogen storage and transport in plants, is synthesized via the ATP-dependent reaction catalyzed by the enzyme asparagine synthetase (AS; EC 6.3.5.4). In this work, we present the molecular analysis of two full-length cDNAs that encode asparagine synthetase in maritime pine (Pinus pinaster Ait.), PpAS1, and PpAS2. Phylogenetic analyses of the deduced amino acid sequences revealed that both genes are class II AS, suggesting an ancient origin of these genes in plants. A comparative study of PpAS1 and PpAS2 gene expression profiles showed that PpAS1 gene is highly regulated by developmental and environmental factors, while PpAS2 is expressed constitutively. To determine the molecular mechanisms underpinning the differential expression of PpAS1, the promoter region of the gene was isolated and putative binding sites for MYB transcription factors were identified. Gel mobility shift assays showed that a MYB protein from Pinus taeda (PtMYB1) was able to interact with the promoter region of PpAS1. Furthermore, transient expression analyses in pine cells revealed a negative effect of PtMYB1 on PpAS1 expression. The potential role of MYB factors in the transcriptional regulation of PpAS1 in vascular cells is discussed. PMID:22654888

  1. Biochemical characterization of the Mycobacterium tuberculosis phosphoribosyl-1-pyrophosphate synthetase

    PubMed Central

    Alderwick, Luke J; Lloyd, Georgina S; Lloyd, Adrian J; Lovering, Andrew L; Eggeling, Lothar; Besra, Gurdyal S

    2011-01-01

    Mycobacterium tuberculosis arabinogalactan (AG) is an essential cell wall component. It provides a molecular framework serving to connect peptidoglycan to the outer mycolic acid layer. The biosynthesis of the arabinan domains of AG and lipoarabinomannan (LAM) occurs via a combination of membrane bound arabinofuranosyltransferases, all of which utilize decaprenol-1-monophosphorabinose as a substrate. The source of arabinose ultimately destined for deposition into cell wall AG or LAM originates exclusively from phosphoribosyl-1-pyrophosphate (pRpp), a central metabolite which is also required for other essential metabolic processes, such as de novo purine and pyrimidine biosyntheses. In M. tuberculosis, a single pRpp synthetase enzyme (Mt-PrsA) is solely responsible for the generation of pRpp, by catalyzing the transfer of pyrophosphate from ATP to the C1 hydroxyl position of ribose-5-phosphate. Here, we report a detailed biochemical and biophysical study of Mt-PrsA, which exhibits the most rapid enzyme kinetics reported for a pRpp synthetase. PMID:21045009

  2. Mammalian folylpoly-. gamma. -glutamate synthetase. 3. Specificity for folate analogues

    SciTech Connect

    George, S.; Cichowicz, D.J.; Shane, B.

    1987-01-27

    A variety of folate analogues were synthesized to explore the specificity of the folate binding site of hog liver folypolyglutamate synthetase and the requirements for catalysis. Modifications of the internal and terminal glutamate moieties of folate cause large drops in on rates and/or affinity for the protein. The only exceptions are glutamine, homocysteate, and ornithine analogues, indicating a less stringent specificity around the delta-carbon of glutamate. It is proposed that initial folate binding to the enzyme involves low-affinity interactions at a pterin and a glutamate site and that the first glutamate bound is the internal residue adjacent to the benzoyl group. Processive movement of the polyglutamate chain through the glutamate site and a possible conformational change in the protein when the terminal residue is bound would result in tight binding and would position the ..gamma..-carboxyl of the terminal glutamate in the correct position for catalysis. The 4-amino substitution of folate increases the on rate for monoglutamate derivatives but severely impairs catalysis with diglutamate derivatives. Pteroylornithine derivatives are the first potent and specific inhibitors of folylpolyglutamate synthetase to be identified and may act as analogues of reaction intermediates. Other folate derivatives with tetrahedral chemistry replacing the peptide bond, such as pteroyl-..gamma..-glutamyl-(psi,CH/sub 2/-NH)-glutamate, retain affinity for the protein but are considerably less effective inhibitors than the ornithine derivatives. Enzyme activity was assayed using (/sup 14/C)glutamate.

  3. MANAGEMENT OF A PATIENT WITH HOLOCARBOXYLASE SYNTHETASE DEFICIENCY

    PubMed Central

    Van Hove, Johan LK; Josefsberg, Sagi; Freehauf, Cynthia; Thomas, Janet A.; Thuy, Le Phuc; Barshop, Bruce A.; Woontner, Michael; Mock, Donald M; Chiang, Pei-Wen; Spector, Elaine; Meneses-Morales, Iván; Cervantes-Roldán, Rafael; León-Del-Río, Alfonso

    2009-01-01

    We investigated in a patient with holocarboxylase synthetase deficiency, the relation between the biochemical and genetic factors of the mutant protein with the pharmacokinetic factors of successful biotin treatment. A girl exhibited abnormal skin at birth, and developed in the first days of life neonatal respiratory distress syndrome and metabolic abnormalities diagnostic of multiple carboxylase deficiency. Enzyme assays showed low carboxylase activities. Fibroblast analysis showed poor incorporation of biotin into the carboxylases, and low transfer of biotin by the holocarboxylase synthetase enzyme. Kinetic studies identified an increased Km but a preserved Vmax. Mutation analysis showed the child to be a compound heterozygote for a new nonsense mutation Q379X and for a novel missense mutation Y663H. This mutation affects a conserved amino acid, which is located the most 3′ of all recorded missense mutations thus far described, and extends the region of functional biotin interaction. Treatment with biotin 100 mg/day gradually improved the biochemical abnormalities in blood and in cerebrospinal fluid, corrected the carboxylase enzyme activities, and provided clinical stability and a normal neurodevelopmental outcome. Plasma concentrations of biotin were increased to more than 500 nM, thus exceeding the increased Km of the mutant enzyme. At these pharmacological concentrations, the CSF biotin concentration was half the concentration in blood. Measuring these pharmacokinetic variables can aid in optimizing treatment, as individual tailoring of dosing to the needs of the mutation may be required. PMID:18974016

  4. Management of a patient with holocarboxylase synthetase deficiency.

    PubMed

    Van Hove, Johan L K; Josefsberg, Sagi; Freehauf, Cynthia; Thomas, Janet A; Thuy, Le Phuc; Barshop, Bruce A; Woontner, Michael; Mock, Donald M; Chiang, Pei-Wen; Spector, Elaine; Meneses-Morales, Iván; Cervantes-Roldán, Rafael; León-Del-Río, Alfonso

    2008-12-01

    We investigated in a patient with holocarboxylase synthetase deficiency, the relation between the biochemical and genetic factors of the mutant protein with the pharmacokinetic factors of successful biotin treatment. A girl exhibited abnormal skin at birth, and developed in the first days of life neonatal respiratory distress syndrome and metabolic abnormalities diagnostic of multiple carboxylase deficiency. Enzyme assays showed low carboxylase activities. Fibroblast analysis showed poor incorporation of biotin into the carboxylases, and low transfer of biotin by the holocarboxylase synthetase enzyme. Kinetic studies identified an increased Km but a preserved Vmax. Mutation analysis showed the child to be a compound heterozygote for a new nonsense mutation Q379X and for a novel missense mutation Y663H. This mutation affects a conserved amino acid, which is located the most 3' of all recorded missense mutations thus far described, and extends the region of functional biotin interaction. Treatment with biotin 100mg/day gradually improved the biochemical abnormalities in blood and in cerebrospinal fluid (CSF), corrected the carboxylase enzyme activities, and provided clinical stability and a normal neurodevelopmental outcome. Plasma concentrations of biotin were increased to more than 500 nM, thus exceeding the increased Km of the mutant enzyme. At these pharmacological concentrations, the CSF biotin concentration was half the concentration in blood. Measuring these pharmacokinetic variables can aid in optimizing treatment, as individual tailoring of dosing to the needs of the mutation may be required.

  5. Purification and properties of the dihydrofolate synthetase from Serratia indica.

    PubMed

    Ikeda, M; Kazuo, I

    1976-01-01

    The dihydrofolate synthetase (EC 6.3.2.12) responsible for catalyzing the synthesis of dihydrofolic acid from dihydropteroic acid and L-glutamic acid was purified about 130-fold from extracts of Serratia indica IFO 3759 by ammonium sulfate fractionation, DEAE-Sephadex column chromatography, Sephadex G-200 gel filtration, and DEAE-cellulose column chromatography. The enzyme preparation obtained was shown to be homogeneous by DEAE-cellulose column chromatography and ultracentrifugal analysis. The sedimentation coefficient of this enzyme was 3.9 S, and the molecular weight was determined to be about 47,000 by Sephadex G-100. The optimum pH for the reaction was 9.0. The enzymatic reaction required dihydropteroate, L-glutamate and ATP as substrates, and Mg2+ and K+ as cofactors. gamma-L-Glutamyl-L-glutamic acid cannot replace L-glutamic acid as the substrate. Neither pteroic acid nor tetrahydropteroic acid can be used as the substrate. ATP was partially replaced by ITP or GTP. The enzyme reaction was inhibited by the addition of AD, but not by AMP. One mole of dihydrofolate, 1 mole of ADP and 1 mole of orthophosphate were produced from each 1 mole of dihydropteroic acid, L-glutamic acid, and ATP by the following equation: 7,8-Dihydropteroic acid ml-Glutamic acid matp Mg2+, K+ leads to Dihydrofolic acid + ADP + Pi. These results suggest that the systematic name for the dihydrofolate synthetase is 7,8-dihydropteroate: L-glutamate ligase (ADP).

  6. Least-Squares Adaptive Control Using Chebyshev Orthogonal Polynomials

    NASA Technical Reports Server (NTRS)

    Nguyen, Nhan T.; Burken, John; Ishihara, Abraham

    2011-01-01

    This paper presents a new adaptive control approach using Chebyshev orthogonal polynomials as basis functions in a least-squares functional approximation. The use of orthogonal basis functions improves the function approximation significantly and enables better convergence of parameter estimates. Flight control simulations demonstrate the effectiveness of the proposed adaptive control approach.

  7. The Gibbs Phenomenon for Series of Orthogonal Polynomials

    ERIC Educational Resources Information Center

    Fay, T. H.; Kloppers, P. Hendrik

    2006-01-01

    This note considers the four classes of orthogonal polynomials--Chebyshev, Hermite, Laguerre, Legendre--and investigates the Gibbs phenomenon at a jump discontinuity for the corresponding orthogonal polynomial series expansions. The perhaps unexpected thing is that the Gibbs constant that arises for each class of polynomials appears to be the same…

  8. The Cytoplasmic Prolyl-tRNA Synthetase of the Malaria Parasite is a Dual-Stage Target for Drug Development

    PubMed Central

    Herman, Jonathan D.; Pepper, Lauren R.; Cortese, Joseph F.; Estiu, Guillermina; Galinsky, Kevin; Zuzarte-Luis, Vanessa; Derbyshire, Emily R.; Ribacke, Ulf; Lukens, Amanda K.; Santos, Sofia A.; Patel, Vishal; Clish, Clary B.; Sullivan, William J.; Zhou, Huihao; Bopp, Selina E.; Schimmel, Paul; Lindquist, Susan; Clardy, Jon; Mota, Maria M.; Keller, Tracy L.; Whitman, Malcolm; Wiest, Olaf; Wirth, Dyann F.; Mazitschek, Ralph

    2015-01-01

    The emergence of drug resistance is a major limitation of current antimalarials. The discovery of new druggable targets and pathways including those that are critical for multiple life cycle stages of the malaria parasite is a major goal for the development of the next-generation of antimalarial drugs. Using an integrated chemogenomics approach that combined drug-resistance selection, whole genome sequencing and an orthogonal yeast model, we demonstrate that the cytoplasmic prolyl-tRNA synthetase (PfcPRS) of the malaria parasite Plasmodium falciparum is a biochemical and functional target of febrifugine and its synthetic derivatives such as halofuginone. Febrifugine is the active principle of a traditional Chinese herbal remedy for malaria. We show that treatment with febrifugine derivatives activated the amino acid starvation response in both P. falciparum and a transgenic yeast strain expressing PfcPRS. We further demonstrate in the P. berghei mouse model of malaria that halofuginol, a new halofuginone analog that we developed, is highly active against both liver and asexual blood stages of the malaria parasite. Halofuginol, unlike halofuginone and febrifugine, is well tolerated at efficacious doses, and represents a promising lead for the development of dual-stage next generation antimalarials. PMID:25995223

  9. Expanding the Genetic Code of Caenorhabditis elegans Using Bacterial aminoacyl-tRNA Synthetase/tRNA Pairs

    PubMed Central

    Parrish, Angela R.; She, Xingyu; Xiang, Zheng; Coin, Irene; Shen, Zhouxin; Briggs, Steven P.; Dillin, Andrew; Wang, Lei

    2012-01-01

    The genetic code specifies 20 common amino acids and is largely preserved in both single and multicellular organisms. Unnatural amino acids (Uaas) have been genetically incorporated into proteins by using engineered orthogonal tRNA/aminoacyltRNA synthetase (RS) pairs, enabling new research capabilities and precision inaccessible with common amino acids. We show here that Escherichia coli tyrosyl and leucyl amber suppressor tRNA/RS pairs can be evolved to incorporate different Uaas in response to the amber stop codon UAG into various proteins in Caenorhabditis elegans. To accurately report Uaa incorporation in worms, we found that it is crucial to integrate the UAG-containing reporter gene into the genome rather than to express it on an extrachromosomal array from which variable expression can lead to reporter activation independent of the amber-suppressing tRNA/RS. Synthesizing a Uaa in a dipeptide drives Uaa uptake and bioavailability. Uaa incorporation has dosage, temporal, tRNA copy, and temperature dependencies similar to endogenous amber suppression. Uaa incorporation efficiency was improved by impairing the nonsense-mediated mRNA decay pathway through knockdown of smg-1. We have generated stable transgenic worms capable of genetically encoding Uaas, enabling Uaa exploitation to address complex biological problems within a metazoan. We anticipate our strategies will be generally extendable to other multicellular organisms. PMID:22554080

  10. Experimental quantum cryptography scheme based on orthogonal states: preliminary results

    NASA Astrophysics Data System (ADS)

    Avella, Alessio; Brida, Giorgio; Degiovanni, Ivo P.; Genovese, Marco; Gramegna, Marco; Traina, Paolo

    2010-04-01

    Since, in general, non-orthogonal states cannot be cloned, any eavesdropping attempt in a Quantum Communication scheme using non-orthogonal states as carriers of information introduces some errors in the transmission, leading to the possibility of detecting the spy. Usually, orthogonal states are not used in Quantum Cryptography schemes since they can be faithfully cloned without altering the transmitted data. Nevertheless, L. Goldberg and L. Vaidman [Phys. Rev. Lett. 75 (7), pp. 12391243, 1995] proposed a protocol in which, even if the data exchange is realized using two orthogonal states, any attempt to eavesdrop is detectable by the legal users. In this scheme the orthogonal states are superpositions of two localized wave packets which travel along separate channels, i.e. two different paths inside a balanced Mach-Zehnder interferometer. Here we present an experiment realizing this scheme.

  11. Anatomy of lithosphere necking during orthogonal rifting

    NASA Astrophysics Data System (ADS)

    Nestola, Yago; Cavozzi, Cristian; Storti, Fabrizio

    2013-04-01

    The evolution of lithosphere necking is a fundamental parameter controlling the structural architecture and thermal-state of rifted margin. The necking shape depends on several parameters, including the extensional strain-rate and thermal layering of the lithosphere. Despite a large number of analogue and numerical modelling studies on lithosphere extension, a quantitative description of the evolution of necking through time is still lacking. We used analogue modelling to simulate in three-dimension the progression of lithosphere thinning and necking during orthogonal rifting. In our models we simulated a typical "cold and young" 4-layer lithosphere stratigraphy: brittle upper crust (loose quartz sand), ductile lower crust (silicon-barite mixture), brittle upper mantle (loose quartz sand), and ductile lower mantle (silicon-barite mixture). The experimental lithosphere rested on a glucose syrup asthenosphere. We monitored model evolution by periodic and coeval laser scanning of both the surface topography and the lithosphere base. After model completion, each of the four layers was removed and the top of the underlying layer was scanned. This technical approach allowed us to quantify the evolution in space and time of the thinning factors for both the whole lithosphere (βz) and the crust (γ). The area of incremental effective stretching (βy) parallel to the extensional direction was obtained from the βz maps.

  12. Nonambipolarity, orthogonal conductivity, poloidal flow, and torque

    SciTech Connect

    Hulbert, G.W.; Perkins, F.W.

    1989-02-01

    Nonambipolar processes, such as neutral injection onto trapped orbits or ripple-diffusion loss of ..cap alpha..-particles, act to charge a plasma. A current j/sub r/ across magnetic surfaces must arise in the bulk plasma to maintain charge neutrality. An axisymmetric, neoclassical model of the bulk plasma shows that these currents are carried by the ions and exert a j/sub r/B/sub theta/R/c torque in the toroidal direction. A driven poloidal flow V/sub theta/ = E/sub r/'c/B must also develop. The average current density is related to the radial electric field E/sub r/' = E/sub r/ + v/sub /phi//B/sub theta//c in a frame moving with the plasma via the orthogonal conductivity = sigma/sub /perpendicular//E/sub r/', which has the value sigma/sub /perpendicular// = (1.65epsilon/sup 1/2/)(ne/sup 2/..nu../sub ii//M..cap omega../sub theta//sup 2/) in the banana regime. If an ignited plasma loses an appreciable fraction ..delta.. of its thermonuclear ..cap alpha..-particles by banana ripple diffusion, then the torque will spin the plasma to sonic rotation in a time /tau//sub s/ approx. 2/tau//sub E//..delta.., /tau//sub E/ being the energy confinement time. 10 refs., 1 fig.

  13. An orthogonal oriented quadrature hexagonal image pyramid

    NASA Technical Reports Server (NTRS)

    Watson, Andrew B.; Ahumada, Albert J., Jr.

    1987-01-01

    An image pyramid has been developed with basis functions that are orthogonal, self-similar, and localized in space, spatial frequency, orientation, and phase. The pyramid operates on a hexagonal sample lattice. The set of seven basis functions consist of three even high-pass kernels, three odd high-pass kernels, and one low-pass kernel. The three even kernels are identified when rotated by 60 or 120 deg, and likewise for the odd. The seven basis functions occupy a point and a hexagon of six nearest neighbors on a hexagonal sample lattice. At the lowest level of the pyramid, the input lattice is the image sample lattice. At each higher level, the input lattice is provided by the low-pass coefficients computed at the previous level. At each level, the output is subsampled in such a way as to yield a new hexagonal lattice with a spacing sq rt 7 larger than the previous level, so that the number of coefficients is reduced by a factor of 7 at each level. The relationship between this image code and the processing architecture of the primate visual cortex is discussed.

  14. Comparison between covariant and orthogonal Lyapunov vectors.

    PubMed

    Yang, Hong-liu; Radons, Günter

    2010-10-01

    Two sets of vectors, covariant Lyapunov vectors (CLVs) and orthogonal Lyapunov vectors (OLVs), are currently used to characterize the linear stability of chaotic systems. A comparison is made to show their similarity and difference, especially with respect to the influence on hydrodynamic Lyapunov modes (HLMs). Our numerical simulations show that in both Hamiltonian and dissipative systems HLMs formerly detected via OLVs survive if CLVs are used instead. Moreover, the previous classification of two universality classes works for CLVs as well, i.e., the dispersion relation is linear for Hamiltonian systems and quadratic for dissipative systems, respectively. The significance of HLMs changes in different ways for Hamiltonian and dissipative systems with the replacement of OLVs with CLVs. For general dissipative systems with nonhyperbolic dynamics the long-wavelength structure in Lyapunov vectors corresponding to near-zero Lyapunov exponents is strongly reduced if CLVs are used instead, whereas for highly hyperbolic dissipative systems the significance of HLMs is nearly identical for CLVs and OLVs. In contrast the HLM significance of Hamiltonian systems is always comparable for CLVs and OLVs irrespective of hyperbolicity. We also find that in Hamiltonian systems different symmetry relations between conjugate pairs are observed for CLVs and OLVs. Especially, CLVs in a conjugate pair are statistically indistinguishable in consequence of the microreversibility of Hamiltonian systems. Transformation properties of Lyapunov exponents, CLVs, and hyperbolicity under changes of coordinate are discussed in appendices.

  15. The Design of the Orthogonal Box Cavity

    SciTech Connect

    Moretti, Alfred; /Fermilab

    2010-09-15

    The muon collider and/or the neutrino factory require large accelerating electric field gradients immersed in large (3 to 6 T) solenoidal magnetic fields for ionization cooling of muon beams. Our original vacuum breakdown study demonstrated a loss of achievable peak accelerating gradient in solenoidal magnetic fields by a factor 2 or greater. The Muon Collaboration has developed a theory of a method to suppress high electric field breakdown in vacuum cavities needed for a Muon collider or neutrino factory. It has been shown in our studies and by others that high gradient electric field emitted electrons (dark current) are the primary cause of breakdown. A DC magnetic field orthogonal to the RF electric accelerating field prevents dark current high field emitted electrons from traveling across the accelerating gap and then will prevent breakdown. We have decided to test this theory by building a special cavity in the shape of vacuum box. Figure 1 is a simplified view of the cavity design. The design is based on an 805 MHz WR975 waveguide cavity resonating in the TE{sub 101} mode. For the TE{sub 101} mode the resonant frequency f{sub 0} is given by the relationship f{sub 0} = c[(I/a){sup 2} + (m/b){sup 2} + (n/d){sup 2}]{sup 0.5}/2 where a and d are the lengths of the base sides and b is the height of the box in MKS units and c is the velocity of light.

  16. Aeroservoelastic modeling with proper orthogonal decomposition

    NASA Astrophysics Data System (ADS)

    Carlson, Henry A.; Verberg, Rolf; Harris, Charles A.

    2017-02-01

    A physics-based, reduced-order, aeroservoelastic model of an F-18 aircraft has been developed using the method of proper orthogonal decomposition (POD), introduced to the field of fluid mechanics by Lumley. The model is constructed with data from high-dimensional, high-fidelity aeroservoelastic computational fluid dynamics (CFD-ASE) simulations that couple equations of motion of the flow to a modal model of the aircraft structure. Through POD modes, the reduced-order model (ROM) predicts both the structural dynamics and the coupled flow dynamics, offering much more information than typically employed, low-dimensional models based on system identification are capable of providing. ROM accuracy is evaluated through direct comparisons between predictions of the flow and structural dynamics with predictions from the parent, the CFD-ASE model. The computational overhead of the ROM is six orders of magnitude lower than that of the CFD-ASE model—accurately predicting the coupled dynamics from simulations of an F-18 fighter aircraft undergoing flutter testing over a wide range of transonic and supersonic flight speeds on a single processor in 1.073 s.

  17. Approximate Orthogonal Sparse Embedding for Dimensionality Reduction.

    PubMed

    Lai, Zhihui; Wong, Wai Keung; Xu, Yong; Yang, Jian; Zhang, David

    2016-04-01

    Locally linear embedding (LLE) is one of the most well-known manifold learning methods. As the representative linear extension of LLE, orthogonal neighborhood preserving projection (ONPP) has attracted widespread attention in the field of dimensionality reduction. In this paper, a unified sparse learning framework is proposed by introducing the sparsity or L1-norm learning, which further extends the LLE-based methods to sparse cases. Theoretical connections between the ONPP and the proposed sparse linear embedding are discovered. The optimal sparse embeddings derived from the proposed framework can be computed by iterating the modified elastic net and singular value decomposition. We also show that the proposed model can be viewed as a general model for sparse linear and nonlinear (kernel) subspace learning. Based on this general model, sparse kernel embedding is also proposed for nonlinear sparse feature extraction. Extensive experiments on five databases demonstrate that the proposed sparse learning framework performs better than the existing subspace learning algorithm, particularly in the cases of small sample sizes.

  18. A reduced basis localized orthogonal decomposition

    NASA Astrophysics Data System (ADS)

    Abdulle, Assyr; Henning, Patrick

    2015-08-01

    In this work we combine the framework of the Reduced Basis method (RB) with the framework of the Localized Orthogonal Decomposition (LOD) in order to solve parametrized elliptic multiscale problems. The idea of the LOD is to split a high dimensional Finite Element space into a low dimensional space with comparably good approximation properties and a remainder space with negligible information. The low dimensional space is spanned by locally supported basis functions associated with the node of a coarse mesh obtained by solving decoupled local problems. However, for parameter dependent multiscale problems, the local basis has to be computed repeatedly for each choice of the parameter. To overcome this issue, we propose an RB approach to compute in an "offline" stage LOD for suitable representative parameters. The online solution of the multiscale problems can then be obtained in a coarse space (thanks to the LOD decomposition) and for an arbitrary value of the parameters (thanks to a suitable "interpolation" of the selected RB). The online RB-LOD has a basis with local support and leads to sparse systems. Applications of the strategy to both linear and nonlinear problems are given.

  19. Analytical modeling of orthogonal spiral structures

    NASA Astrophysics Data System (ADS)

    Santos, Auteliano A.; Hobeck, Jared D.; Inman, Daniel J.

    2016-11-01

    This paper presents the analytical modeling of orthogonal spiral structures (OSS), a promising option for small-scale energy harvesting applications. This unique multi-beam structure is analyzed using a distributed parameter approach with Euler-Bernoulli assumptions. First, an aluminum substrate is evaluated to determine if the proposed design can be used to capture vibration energy in the desired frequency range using a twelve beam OSS. Finite element calculations are used to validate the analytical model. This model is then modified to include the electromechanical effects of a piezoelectric layer added to the aluminum substrate. Lastly, the effects of the beam width and the number of beams is analyzed for a particular surface area of the OSS. Results show that increasing the number of beams causes a reduction in the first natural frequency. From those results, it is possible to conclude that OSS can be used as an alternative to current energy harvesting systems for MEMS applications, allowing the capture of environmental energy in the frequency range of common mechanical systems.

  20. Effect of Liver Damage and Hyperbaric Oxygenation on Glutamine Synthetase of Hepatocytes.

    PubMed

    Savilov, P N; Yakovlev, V N

    2016-01-01

    Activity of glutamine synthetase in the hepatocytes of healthy animals and animals with chronic CCl4-induced hepatitis was studied on white mature female rats after liver resection (15-20% of organ weight) and hyperbaric oxygenation (3 atm, 50 min, 3 times). Surgically operated left and non-operated middle lobes of the liver were analyzed on day 3 after liver resection and exposure to hyperbaric oxygenation. On day 65 of CCl4 poisoning, activity of glutamine synthetase decreased in both lobes and did not recover on day 3 after toxin cessation. Liver resection under conditions of CCl4-induced hepatitis restored reduced activity of glutamine synthetase in both liver lobes to the normal level. In healthy rats, the increase in glutamine synthetase activity after liver resection was found only in the middle lobe of the liver. Hyperbaric oxygenation enhanced the stimulatory effect of liver resection on glutamine synthetase activity in hepatocytes during chronic CCl4-induced hepatitis. In healthy animals with liver resection, activity of glutamine synthetase did not change after hyperbaric oxygenation, while normally oxygenation inhibited glutamine synthetase activity.

  1. Encapsulation of glutamine synthetase in mouse erythrocytes: a new procedure for ammonia detoxification.

    PubMed

    Kosenko, Elena A; Venediktova, Natalia I; Kudryavtsev, Andrey A; Ataullakhanov, Fazoil I; Kaminsky, Yury G; Felipo, Vicente; Montoliu, Carmina

    2008-12-01

    There are a number of pathological situations in which ammonia levels increase leading to hyperammonemia, which may cause neurological alterations and can lead to coma and death. Currently, there are no efficient treatments allowing rapid and sustained decrease of ammonia levels in these situations. A way to increase ammonia detoxification would be to increase its incorporation in glutamine by glutamine synthetase. The aim of this work was to develop a procedure to encapsulate glutamine synthetase in mouse erythrocytes and to assess whether administration of these erythrocytes containing glutamine synthetase (GS) reduce ammonia levels in hyperammonemic mice. The procedure developed allowed the encapsulation of 3 +/- 0.25 IU of GS / mL of erythrocytes with a 70% cell recovery. Most metabolites, including ATP, remained unaltered in glutamine synthetase-loaded erythrocytes (named ammocytes by us) compared with native erythrocytes. The glutamine synthetase-loaded ammocytes injected in mice survived and retained essentially all of their glutamine synthetase activity for at least 48 h in vivo. Injection of these ammocytes into hyperammonemic mice reduced ammonia levels in the blood by about 50%. The results reported indicate that ammocytes are able to keep their integrity, normal energy metabolism, the inserted glutamine synthetase activity, and can be useful to reduce ammonia levels in hyperammonemic situations.

  2. Molecular structure of the human argininosuccinate synthetase gene: Occurrence of alternative mRNA splicing

    SciTech Connect

    Freytag, S.O.; Beaudet, A.L.; Bock, H.G.O.; O'Brien, W.E.

    1984-10-01

    The human genome contains one expressed argininosuccinate synthetase gene and ca. 14 pseudogenes that are dispersed to at least 11 human chromosomes. Eleven clones isolated from a human genomic DNA library were characterized extensively by restriction mapping, Southern blotting, and nucleotide sequencing. These 11 clones represent the entire expressed argininosuccinate synthetase gene that spans 63 kilobases and contains at least 13 exons. The expressed gene codes for two mRNAs that differ in their 5' untranslated sequences and arise by alternative splicing involving the inclusion or deletion of an entire exon. In normal human liver and cultured fibroblasts, the predominant mature argininosuccinate synthetase mRNA lacks sequences encoded by exon 2 in the expressed gene. In contrast, the predominant argininosuccinate synthetase mRNA in baboon liver contains exon 2 sequences. A transformed canavanine-resistant human cell line in which argininosuccinate synthetase activity is 180-fold higher than that in wild-type cells contains abundant amounts of both forms of the argininosuccinate synthetase mRNA. The mRNA lacking exon 2 sequences is the more abundant mRNA species in the canavanine-resistant cells. These observations show that splicing of the argininosuccinate synthetase mRNA is species specific in primates and varies among different human cell types.

  3. Computing the roots of complex orthogonal and kernel polynomials

    SciTech Connect

    Saylor, P.E.; Smolarski, D.C.

    1988-01-01

    A method is presented to compute the roots of complex orthogonal and kernel polynomials. An important application of complex kernel polynomials is the acceleration of iterative methods for the solution of nonsymmetric linear equations. In the real case, the roots of orthogonal polynomials coincide with the eigenvalues of the Jacobi matrix, a symmetric tridiagonal matrix obtained from the defining three-term recurrence relationship for the orthogonal polynomials. In the real case kernel polynomials are orthogonal. The Stieltjes procedure is an algorithm to compute the roots of orthogonal and kernel polynomials bases on these facts. In the complex case, the Jacobi matrix generalizes to a Hessenberg matrix, the eigenvalues of which are roots of either orthogonal or kernel polynomials. The resulting algorithm generalizes the Stieljes procedure. It may not be defined in the case of kernel polynomials, a consequence of the fact that they are orthogonal with respect to a nonpositive bilinear form. (Another consequence is that kernel polynomials need not be of exact degree.) A second algorithm that is always defined is presented for kernel polynomials. Numerical examples are described.

  4. Critical Evaluation of the Changes in Glutamine Synthetase Activity in Models of Cerebral Stroke.

    PubMed

    Jeitner, Thomas M; Battaile, Kevin; Cooper, Arthur J L

    2015-12-01

    The following article addresses some seemingly paradoxical observations concerning cerebral glutamine synthetase in ischemia-reperfusion injury. In the brain, this enzyme is predominantly found in astrocytes and catalyzes part of the glutamine-glutamate cycle. Glutamine synthetase is also thought to be especially sensitive to inactivation by the oxygen- and nitrogen-centered radicals generated during strokes. Despite this apparent sensitivity, glutamine synthetase specific activity is elevated in the affected tissues during reperfusion. Given the central role of the glutamine-glutamate cycle in the brain, we sought to resolve these conflicting observations with the view of providing an alternative perspective for therapeutic intervention in stroke.

  5. Isolation of a cDNA clone for human threonyl-tRNA synthetase

    SciTech Connect

    Kontis, K.J.; Arfin, S.M.

    1989-05-01

    A cDNA for threonyl-tRNA synthetase was isolated from a human placental cDNA /lambda/gt11 expression library by immunological screening, and its identity was confirmed by hybrid-selected mRNA translation. With this cDNA used as a hybridization probe, borrelidin-resistant Chinese hamster ovary cells that overproduced threonyl-tRNA synthetase were shown to have increased levels of threonyl-tRNA synthetase mRNA and gene sequences. Amplification of the gene did not appear to have been accompanied by any major structural reorganizations.

  6. Organizing pneumonia as the first manifestation of anti-synthetase syndrome.

    PubMed

    Priyangika, S M Thanuja Nilushi; Karunarathna, W G S G; Liyanage, Isurujith; Gunawardana, Methsala; Udumalgala, Sumeda; Rosa, Chamith; Kulatunga, Aruna

    2016-06-02

    Anti-synthetase syndrome associated interstitial lung disease can occur either simultaneously, before, or after the development of polymyositis/dermatomyositis. Histology of interstitial lung disease can be nonspecific interstitial pneumonia, usual interstitial pneumonia, diffuse alveolar damage, organizing pneumonia. Organizing pneumonia associated anti-synthetase syndrome is a rare finding especially as the first manifestation. We report a 41 year old male patient who presented with organizing pneumonia and 2 years following the onset, developed polymyositis with anti-JO-1 antibody positivity. It is important to screen patients with organizing pneumonia for anti-synthetase syndrome which can be manifested later.

  7. The MTCY428.08 Gene of Mycobacterium tuberculosis Codes for NAD+ Synthetase

    PubMed Central

    Cantoni, Rita; Branzoni, Manuela; Labò, Monica; Rizzi, Menico; Riccardi, Giovanna

    1998-01-01

    The product of the MTCY428.08 gene of Mycobacterium tuberculosis shows sequence homology with several NAD+ synthetases. The MTCY428.08 gene was cloned into the expression vectors pGEX-4T-1 and pET-15b. Expression in Escherichia coli led to overproduction of glutathione S-transferase fused and His6-tagged gene products, which were enzymatically assayed for NAD synthetase activity. Our results demonstrate that the MTCY428.08 gene of M. tuberculosis is the structural gene for NAD+ synthetase. PMID:9620974

  8. Heterogeneity of holocarboxylase synthetase in patients with biotin-responsive multiple carboxylase deficiency.

    PubMed Central

    Burri, B J; Sweetman, L; Nyhan, W L

    1985-01-01

    Holocarboxylase synthetase activity has been determined in fibroblasts of seven patients with the neonatal form of biotin-responsive multiple carboxylase deficiency. The normal Km for biotin was 15 +/- 3 nmol/l, while in the patients the values ranged from 48 to 1,062 nmol/l. The mean maximum velocity was 27% of normal. Differences among the values obtained for the Km for biotin and the heat stability of holocarboxylase synthetase suggested that the patients studied represented at least four distinct variants at the holocarboxylase synthetase locus. PMID:3920902

  9. Revised nomenclature for the mammalian long-chain acyl-CoA synthetase gene family.

    PubMed

    Mashek, Douglas G; Bornfeldt, Karin E; Coleman, Rosalind A; Berger, Johannes; Bernlohr, David A; Black, Paul; DiRusso, Concetta C; Farber, Steven A; Guo, Wen; Hashimoto, Naohiro; Khodiyar, Varsha; Kuypers, Frans A; Maltais, Lois J; Nebert, Daniel W; Renieri, Alessandra; Schaffer, Jean E; Stahl, Andreas; Watkins, Paul A; Vasiliou, Vasilis; Yamamoto, Tokuo T

    2004-10-01

    By consensus, the acyl-CoA synthetase (ACS) community, with the advice of the human and mouse genome nomenclature committees, has revised the nomenclature for the mammalian long-chain acyl-CoA synthetases. ACS is the family root name, and the human and mouse genes for the long-chain ACSs are termed ACSL1,3-6 and Acsl1,3-6, respectively. Splice variants of ACSL3, -4, -5, and -6 are cataloged. Suggestions for naming other family members and for the nonmammalian acyl-CoA synthetases are made.

  10. Polar Plate Theory for Orthogonal Anisotropy

    NASA Technical Reports Server (NTRS)

    Bailey, Michelle D.; Bower, Mark V.

    2000-01-01

    Laminated fiber-reinforced (or filamentary) composites are used today for their high strength-to-weight and stiffness-to-weight ratios. However, because of the anisotropic behavior of composites, determining the response on a macroscopic scale is challenging. This is particularly evident in the evaluation of the governing differential equations of a circular disk with the fibers of the lamina oriented with rectilinear orthogonality. This includes any situation involving a composite plate of circular geometry in which out-of-plane displacements due to load are desired, such as fastener pull through loading of a composite plate. Current analysis techniques use numerical methods with rectilinear coordinate systems to solve problems with circular geometry. These analyses over predict plate stiffness by 20% and underpredict failure by 70%. Consequently, there is a need to transform classical composite plate theory to a polar coordinate system. In order to better analyze structures with circular geometries the classical composite plate equations are transformed into the plate equations for a rectilinearly anisotropic composite in polar coordinates. A composite plate is typically a laminate of fibers in rectilinear directions. Subsequent to the lay-tip the necessary geometry is cut out of a rectangular plate. In a similar manner, the derivation of the plate equation starts with the fundamental definitions of strain, displacement and curvature and incorporates the material property angular dependence into the equilibrium equations for a differential polar element. In the transformed state, the stiffness coefficients are no longer constant, adding to the complexity of the governing differential equations. This paper discusses the new derivation and evaluation of the plate equations for a circular composite disk with orthogonal rectilinear anisotropy. The resultant new three partial differential equations, which describe the circular anisotropic plate, can be used to

  11. Subjective ranking of concert halls substantiated through orthogonal objective parameters.

    PubMed

    Cerdá, Salvador; Giménez, Alicia; Cibrián, Rosa; Girón, Sara; Zamarreño, Teófilo

    2015-02-01

    This paper studies the global subjective assessment, obtained from mean values of the results of surveys addressed to members of the audience of live concerts in Spanish auditoriums, through the mean values of the three orthogonal objective parameters (Tmid, IACCE3, and LEV), expressed in just noticeable differences (JNDs), regarding the best-valued hall. Results show that a linear combination of the relative variations of orthogonal parameters can largely explain the overall perceived quality of the sample. However, the mean values of certain orthogonal parameters are not representative, which shows that an alternative approach to the problem is necessary. Various possibilities are proposed.

  12. Aminoacyl-tRNA synthetases in medicine and disease

    PubMed Central

    Yao, Peng; Fox, Paul L

    2013-01-01

    Aminoacyl-tRNA synthetases (ARSs) are essential and ubiquitous ‘house-keeping’ enzymes responsible for charging amino acids to their cognate tRNAs and providing the substrates for global protein synthesis. Recent studies have revealed a role of multiple ARSs in pathology, and their potential use as pharmacological targets and therapeutic reagents. The ongoing discovery of genetic mutations in human ARSs is increasing exponentially and can be considered an important determinant of disease etiology. Several chemical compounds target bacterial, fungal and human ARSs as antibiotics or disease-targeting medicines. Remarkably, ongoing exploration of noncanonical functions of ARSs has shown important contributions to control of angiogenesis, inflammation, tumourigenesis and other important physiopathological processes. Here, we summarize the roles of ARSs in human diseases and medicine, focusing on the most recent and exciting discoveries. PMID:23427196

  13. Regulation of Angiogenesis by Aminoacyl-tRNA Synthetases

    PubMed Central

    Mirando, Adam C.; Francklyn, Christopher S.; Lounsbury, Karen M.

    2014-01-01

    In addition to their canonical roles in translation the aminoacyl-tRNA synthetases (ARSs) have developed secondary functions over the course of evolution. Many of these activities are associated with cellular survival and nutritional stress responses essential for homeostatic processes in higher eukaryotes. In particular, six ARSs and one associated factor have documented functions in angiogenesis. However, despite their connection to this process, the ARSs are mechanistically distinct and exhibit a range of positive or negative effects on aspects of endothelial cell migration, proliferation, and survival. This variability is achieved through the appearance of appended domains and interplay with inflammatory pathways not found in prokaryotic systems. Complete knowledge of the non-canonical functions of ARSs is necessary to understand the mechanisms underlying the physiological regulation of angiogenesis. PMID:25535072

  14. Biochemical and structural investigations on phosphoribosylpyrophosphate synthetase from Mycobacterium smegmatis

    PubMed Central

    Donini, Stefano; Garavaglia, Silvia; Ferraris, Davide M.; Miggiano, Riccardo; Mori, Shigetarou; Shibayama, Keigo

    2017-01-01

    Mycobacterium smegmatis represents one model for studying the biology of its pathogenic relative Mycobacterium tuberculosis. The structural characterization of a M. tuberculosis ortholog protein can serve as a valid tool for the development of molecules active against the M. tuberculosis target. In this context, we report the biochemical and structural characterization of M. smegmatis phosphoribosylpyrophosphate synthetase (PrsA), the ortholog of M. tuberculosis PrsA, the unique enzyme responsible for the synthesis of phosphoribosylpyrophosphate (PRPP). PRPP is a key metabolite involved in several biosynthetic pathways including those for histidine, tryptophan, nucleotides and decaprenylphosphoryl-arabinose, an essential precursor for the mycobacterial cell wall biosynthesis. Since M. tuberculosis PrsA has been validated as a drug target for the development of antitubercular agents, the data presented here will add to the knowledge of the mycobacterial enzyme and could contribute to the development of M. tuberculosis PrsA inhibitors of potential pharmacological interest. PMID:28419153

  15. Redesigning the stereospecificity of tyrosyl-tRNA synthetase.

    PubMed

    Simonson, Thomas; Ye-Lehmann, Shixin; Palmai, Zoltan; Amara, Najette; Wydau-Dematteis, Sandra; Bigan, Erwan; Druart, Karen; Moch, Clara; Plateau, Pierre

    2016-02-01

    D-Amino acids are largely excluded from protein synthesis, yet they are of great interest in biotechnology. Unnatural amino acids have been introduced into proteins using engineered aminoacyl-tRNA synthetases (aaRSs), and this strategy might be applicable to D-amino acids. Several aaRSs can aminoacylate their tRNA with a D-amino acid; of these, tyrosyl-tRNA synthetase (TyrRS) has the weakest stereospecificity. We use computational protein design to suggest active site mutations in Escherichia coli TyrRS that could increase its D-Tyr binding further, relative to L-Tyr. The mutations selected all modify one or more sidechain charges in the Tyr binding pocket. We test their effect by probing the aminoacyl-adenylation reaction through pyrophosphate exchange experiments. We also perform extensive alchemical free energy simulations to obtain L-Tyr/D-Tyr binding free energy differences. Agreement with experiment is good, validating the structural models and detailed thermodynamic predictions the simulations provide. The TyrRS stereospecificity proves hard to engineer through charge-altering mutations in the first and second coordination shells of the Tyr ammonium group. Of six mutants tested, two are active towards D-Tyr; one of these has an inverted stereospecificity, with a large preference for D-Tyr. However, its activity is low. Evidently, the TyrRS stereospecificity is robust towards charge rearrangements near the ligand. Future design may have to consider more distant and/or electrically neutral target mutations, and possibly design for binding of the transition state, whose structure however can only be modeled.

  16. Transformation of Bacillus Subtilis with cloned thymidylate synthetases

    SciTech Connect

    Rubin, Edward M.

    1980-01-01

    Bacillus subtilis carries two genes, thyA and thyB, each encoding different protein products, with thymidylate synthetase (TSase) activity. Either of these genes alone is sufficient for thymidine independence in B. subtilis. In addition there exist two B. subtilis temperate bacteriophages which upon infection of thymine requiring auxotrophs results in conversion of the organism to thymine independence. Chimeric plasmids selected for Thy/sup +/ transforming activity in E. coli were constructed and then used as a source of defined highly enriched DNA with which to transform competent B. subtilis. These plasmids were studied for their: (1) abiility to transform B. subtilis to thymine independence; (2) site of integration within the B. subtilis chromosome upon transformation; (3) phenotype of Thy/sup +/ plasmid generated transformants; and (4) nucleotide sequence homology among the cloned DNA fragments conferring thymine independence. Plasmids containing the two bacteriophage thy genes displayed the phenotype associated with thyA, whereas the plasmids containing the cloned B. subtilis chromosomal genes displayed the phenotype associated with thyB. Utilizing similar technology, the ability of an entirely foreign hybred bacterial plasmiid to transform B. subtilis was examined. In this case the gene from E. coli encoding thymidylate synthetase was cloned in the plasmid pBR322. The resulting chimeric plasmid was effective in transforming both E. coli and B. subtilis to thymine prototrophy. Uncloned linear E. coli chromosomal DNA was unable to transform thymine requiring strains of B. subtilis to thymine independence. Although the Thy/sup +/ transformants of E. coli contained plasmid DNA, the Thy/sup +/ transformants derived from the transformation of B. subtilis did not contain detectable extrachromosomal DNA. Instead the DNA from the chimeric plasmid was integrated into the chromosome of B. subtilis. (ERB)

  17. Activity of formylphosphate in the reaction catalyzed by formyltetrahydrofolate synthetase

    SciTech Connect

    Jahansouz, H.; Kofron, J.L.; Smithers, G.W.; Himes, R.H.; Reed, G.H.

    1986-05-01

    Formylphosphate (FP), a putative enzyme-bound intermediate in the reaction catalyzed by N/sup 10/-formylH/sub 4/folate synthetase, was synthesized from formylfluoride and Pi. Measurement of hydrolysis rates by /sup 31/P NMR showed that FP is very unstable with a half-life of 48 min at 20/sup 0/C and pH 7. At pH 7 hydrolysis occurs with O-P bond cleavage as shown by /sup 18/O incorporation from /sup 18/O-H/sub 2/O into Pi. The substrate activity of FP was tested in the reaction catalyzed by N/sup 10/-formylH/sub 4/folate synthetase isolated from Clostridium cylindrosporum. MgATP + H/sub 4/folate + HCOO/sup -/ in equilibrium MgADP + Pi +N/sup 10/-formylH/sub 4/folate FP supports the reaction in both the forward and reverse directions. Thus, N/sup 10/-formylH/sub 4/folate is produced from H/sub 4/-folate and FP but only if ADP is present, and ATP is produced from FP and ADP but only if H/sub 4/folate is present. The requirements for H/sub 4/folate in the synthesis of ATP from ADP and FP and for ADP in the synthesis of N/sup 10/-formylH/sub 4/folate from FP and H/sub 4/folate, are consistent with past kinetic and isotope exchange studies which showed that the reaction proceeds by a sequential mechanism and that all three substrates must be present for any reaction to occur.

  18. Identification of pantoate kinase and phosphopantothenate synthetase from Methanospirillum hungatei.

    PubMed

    Katoh, Hiroki; Tamaki, Hideyuki; Tokutake, Yuka; Hanada, Satoshi; Chohnan, Shigeru

    2013-04-01

    Pantothenate synthetase (PanC) and pantothenate kinase which function in the canonical coenzyme A (CoA) biosynthetic pathway cannot be found in most archaea. COG1829 and COG1701 intrinsic to archaea were proposed as the candidate proteins for producing 4'-phosphopantothenate instead, and the COG1701 protein from Methanosarcina mazei was assigned as PanC. Meanwhile, the Thermococcus kodakarensis COG1829 and COG1701 proteins were biochemically identified as novel enzymes, i.e., pantoate kinase (PoK) and phosphopantothenate synthetase (PPS). In this study, the functions of Mhun_0831 (COG1829) and Mhun_0832 (COG1701) from Methanospirillum hungatei were identified, and the recombinant enzymes were partially characterized. Plasmids simultaneously possessing the two genes encoding Mhun_0831 and Mhun_0832 complemented the poor growth of the temperature-sensitive Escherichia coli pantothenate kinase mutant ts9. The recombinant Mhun_0831 and Mhun_0832 expressed in E. coli cells exhibited PoK and PPS activities, respectively, being in accord with the functions of T. kodakarensis proteins. The PoK activity was most active at pH 8.5 and 40°C, and accepted ATP and UTP as a phosphate donor. Although CoA did not affect the PoK activity, the end product considerably accelerated the PPS activity. The homologs of both proteins are widely conserved in most archaeal genomes. Taken together, our findings indicate that archaea can synthesize CoA through the unique pathway involving PoK and PPS, in addition to the canonical one that the order Thermoplasmatales employs.

  19. Identification and functional characterization of a novel bacterial type asparagine synthetase A: a tRNA synthetase paralog from Leishmania donovani.

    PubMed

    Manhas, Reetika; Tripathi, Pankaj; Khan, Sameena; Sethu Lakshmi, Bhavana; Lal, Shambhu Krishan; Gowri, Venkatraman Subramanian; Sharma, Amit; Madhubala, Rentala

    2014-04-25

    Asparagine is formed by two structurally distinct asparagine synthetases in prokaryotes. One is the ammonia-utilizing asparagine synthetase A (AsnA), and the other is asparagine synthetase B (AsnB) that uses glutamine or ammonia as a nitrogen source. In a previous investigation using sequence-based analysis, we had shown that Leishmania spp. possess asparagine-tRNA synthetase paralog asparagine synthetase A (LdASNA) that is ammonia-dependent. Here, we report the cloning, expression, and kinetic analysis of ASNA from Leishmania donovani. Interestingly, LdASNA was both ammonia- and glutamine-dependent. To study the physiological role of ASNA in Leishmania, gene deletion mutations were attempted via targeted gene replacement. Gene deletion of LdASNA showed a growth delay in mutants. However, chromosomal null mutants of LdASNA could not be obtained as the double transfectant mutants showed aneuploidy. These data suggest that LdASNA is essential for survival of the Leishmania parasite. LdASNA enzyme was recalcitrant toward crystallization so we instead crystallized and solved the atomic structure of its close homolog from Trypanosoma brucei (TbASNA) at 2.2 Å. A very significant conservation in active site residues is observed between TbASNA and Escherichia coli AsnA. It is evident that the absence of an LdASNA homolog from humans and its essentiality for the parasites make LdASNA a novel drug target.

  20. Fast Orthogonal Row-Column Electronic Scanning With Top-Orthogonal-to-Bottom Electrode Arrays.

    PubMed

    Ceroici, Chris; Harrison, Tyler; Zemp, Roger J

    2017-06-01

    Recently, top-orthogonal-to-bottom electrode 2-D arrays were introduced as a practical design for 3-D ultrasound imaging without requiring the wiring of a 2-D grid of elements. However, previously proposed imaging schemes suffered from speed or image-quality limitations. Here, we propose a new imaging scheme which we call Fast Orthogonal Row-Column Electronic Scanning (FORCES). This new approach takes advantage of bias sensitivity to enable high-quality and fast B-scan imaging. We compare this imaging scheme with an equivalent linear array, a previously proposed row-column imaging scheme, as well as with the Explososcan imaging scheme for 2-D arrays through simulations. In a point phantom simulation, the lateral (azimuthal) resolution of a 64 ×64 element 6.67-MHz λ /2-pitch array using the FORCES imaging scheme with an f-number of 1.7 was 0.52 mm with similar in-plane image quality to an equivalent linear array but with improved and electronically steerable elevational resolution. When compared with other 3-D imaging schemes in point phantom simulations, the FORCES imaging scheme showed an azimuthal resolution improvement of 54% compared with Explososcan. Compared with a previously introduced row-column method, the FORCES imaging scheme had similar resolution but a 25-dB decrease in sidelobe amplitude, significantly impacting contrast to noise in scattering phantoms.

  1. Nonlocality of orthogonal product-basis quantum states

    NASA Astrophysics Data System (ADS)

    Wang, Yan-Ling; Li, Mao-Sheng; Zheng, Zhu-Jun; Fei, Shao-Ming

    2015-09-01

    We study the local indistinguishability of mutually orthogonal product basis quantum states in the high-dimensional quantum system. In the quantum system of Cd⊗Cd , where d is odd, Zhang et al. [Z.-C. Zhang et al., Phys. Rev. A 90, 022313 (2014), 10.1103/PhysRevA.90.022313] have constructed d2 orthogonal product basis quantum states that are locally indistinguishable. We find a subset that contains 6 d -9 orthogonal product states that are still locally indistinguishable. We generalize our method to an arbitrary bipartite quantum system Cm⊗Cn . We present a small set with only 3 (m +n )-9 orthogonal product states and prove that these states are local operations and classical communication (LOCC) indistinguishable. Even though these 3 (m +n )-9 product states are LOCC indistinguishable, they can be distinguished by separable measurements. This shows that separable operations are strictly stronger than the local operations and classical communication.

  2. 13. Credit JTL: Detail, orthogonal view of Egyptian Revivial decorative ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    13. Credit JTL: Detail, orthogonal view of Egyptian Revivial decorative motifs used typically at midpoints of diagonals - Reading-Halls Station Bridge, U.S. Route 220, spanning railroad near Halls Station, Muncy, Lycoming County, PA

  3. Systems of Differential Equations with Skew-Symmetric, Orthogonal Matrices

    ERIC Educational Resources Information Center

    Glaister, P.

    2008-01-01

    The solution of a system of linear, inhomogeneous differential equations is discussed. The particular class considered is where the coefficient matrix is skew-symmetric and orthogonal, and where the forcing terms are sinusoidal. More general matrices are also considered.

  4. Systems of Differential Equations with Skew-Symmetric, Orthogonal Matrices

    ERIC Educational Resources Information Center

    Glaister, P.

    2008-01-01

    The solution of a system of linear, inhomogeneous differential equations is discussed. The particular class considered is where the coefficient matrix is skew-symmetric and orthogonal, and where the forcing terms are sinusoidal. More general matrices are also considered.

  5. Generalized Rayleigh and Jacobi Processes and Exceptional Orthogonal Polynomials

    NASA Astrophysics Data System (ADS)

    Chou, C.-I.; Ho, C.-L.

    2013-09-01

    We present four types of infinitely many exactly solvable Fokker-Planck equations, which are related to the newly discovered exceptional orthogonal polynomials. They represent the deformed versions of the Rayleigh process and the Jacobi process.

  6. Self-mixing interferometry with mutual independent orthogonal polarized light.

    PubMed

    Zhang, Shaohui; Zhang, Shulian; Tan, Yidong; Sun, Liqun

    2016-02-15

    A self-mixing interferometry with mutual independent orthogonal polarized light is introduced. Its most important feature is that two mutual independent orthogonal lights are used as measuring and reference light. Frequency shifting and polarization multiplexing technologies are used in the proposed optical system. Phase variation of the two orthogonal polarized beams is simultaneously measured through heterodyne demodulation with a lock-in amplifier. The phase difference of the orthogonal polarized light accurately reflects the target displacement. The target in this system is a non-cooperative object which is different from a traditional Michelson interferometer. The primary experimental results show that this kind of self-mixing interferometry is very feasible. Under typical room conditions, the system's short-term resolution is better than 2 nm.

  7. Purification and determination of glutamine synthetase by high-performance immunoaffinity chromatography.

    PubMed

    Alhama, J; López-Barea, J; Toribio, F; Roldán, J M

    1992-01-10

    High-performance immunoaffinity chromatography (HPIAC) with anti-glutamine synthetase polyclonal antibodies bound to epoxy-activated silica was used to purify and determine this enzyme from the cyanobacterium Synechocystis. A single-step HPIAC procedure with cell-free extracts yielded electroporetically homogeneous glutamine synthetase. In the determination of glutamine synthetase by HPIAC a linear response in the range 10-60 micrograms of enzyme was observed. Recoveries of 70% of the loaded enzymatic activity and 100% of protein were obtained. The determination of glutamine synthetase protein by HPIAC was compared with that obtained by rocket immunoelectrophoresis. The chromatographic method is proposed as a possible alternative to other immunochemical quantitative techniques, particularly when non-limiting amounts of samples are available.

  8. Preparation and cross-reactivity of anti-avian glutamine synthetase antibody.

    PubMed

    Smith, D D; Vorhaben, J E; Campbell, J W

    1983-04-01

    Rabbit antibody to chicken liver mitochondrial glutamine synthetase was purified by immunoaffinity chromatography for analysis of the immunological relatedness of vertebrate glutamine synthetases. The antibody cross-reacted with enzymes from representatives of all five vertebrate classes, indicating a high degree of evolutionary conservatism in the structure of the enzymes. A unique aspect of the immunological similarity of these enzymes is that it exists between cytosolic and mitochondrial enzymes which are, in general, immunologically distinct. The antibody did not cross-react with two insect glutamine synthetases. Compositional difference indices, calculated from the amino acid compositions of glutamine synthetases from several species, gave a mean estimate of over 80% sequence homology for the vertebrate enzymes. The avian mitochondrial enzyme gave a mean 78% homology with the mammalian cytosolic enzyme.

  9. Evidence for two immunologically distinct acetyl-coenzyme A synthetases in yeast

    NASA Technical Reports Server (NTRS)

    Satyanarayana, T.; Mandel, A. D.; Klein, H. P.

    1974-01-01

    Evidence is presented that clearly establishes the presence of two acetyl-CoA synthetases in Saccharomyces cerevisiae, one elaborated under 'aerobic' conditions, the other under 'nonaerobic' conditions. The antibody produced by each enzyme is immunologically specific.

  10. Regulation of active site coupling in glutamine-dependent NAD[superscript +] synthetase

    SciTech Connect

    LaRonde-LeBlanc, Nicole; Resto, Melissa; Gerratana, Barbara

    2009-05-21

    NAD{sup +} is an essential metabolite both as a cofactor in energy metabolism and redox homeostasis and as a regulator of cellular processes. In contrast to humans, Mycobacterium tuberculosis NAD{sup +} biosynthesis is absolutely dependent on the activity of a multifunctional glutamine-dependent NAD{sup +} synthetase, which catalyzes the ATP-dependent formation of NAD{sup +} at the synthetase domain using ammonia derived from L-glutamine in the glutaminase domain. Here we report the kinetics and structural characterization of M. tuberculosis NAD{sup +} synthetase. The kinetics data strongly suggest tightly coupled regulation of the catalytic activities. The structure, the first of a glutamine-dependent NAD{sup +} synthetase, reveals a homooctameric subunit organization suggesting a tight dependence of catalysis on the quaternary structure, a 40-{angstrom} intersubunit ammonia tunnel and structural elements that may be involved in the transfer of information between catalytic sites.

  11. Severe holocarboxylase synthetase deficiency with incomplete biotin responsiveness resulting in antenatal insult in samoan neonates.

    PubMed

    Wilson, Callum J; Myer, Michael; Darlow, Brian A; Stanley, Thorsten; Thomson, Glen; Baumgartner, E Regula; Kirby, Denise M; Thorburn, David R

    2005-07-01

    We describe 7 Polynesian babies with a unique severe form of holocarboxylase synthetase deficiency characterized by antenatal growth retardation, subependymal cysts, only partial response to biotin, and a poor outcome.

  12. Alternative pathways for editing non-cognate amino acids by aminoacyl-tRNA synthetases.

    PubMed Central

    Jakubowski, H; Fersht, A R

    1981-01-01

    Evidence is presented that the editing mechanisms of aminoacyl-tRNA synthetase operate by two alternative pathways: pre-transfer, by hydrolysis of the non-cognate aminoacyl adenylate; post-transfer, by hydrolysis of the mischarged tRNA. The methionyl-tRNA synthetases from Escherichia coli and Bacillus stearothermophilus and isoleucyl-tRNA synthetase from E. coli, for example, are shown to reject misactivated homocysteine rapidly by the pre-transfer route. A novel feature of this reaction is that homocysteine thiolactone is formed by the facile cyclisation of the homocysteinyl adenylate. Valyl-tRNA synthetases, on the other hand, reject the more readily activated non-cognate amino acids by primarily the post-transfer route. The features governing the choice of pathway are discussed. PMID:7024910

  13. Functional expansion of human tRNA synthetases achieved by structural inventions

    PubMed Central

    Guo, Min; Schimmel, Paul; Yang, Xiang-Lei

    2010-01-01

    Known as an essential component of the translational apparatus, the aminoacyl-tRNA synthetase family catalyzes the first step reaction in protein synthesis, that is, to specifically attach each amino acid to its cognate tRNA. While preserving this essential role, tRNA synthetases developed other roles during evolution. Human tRNA synthetases, in particular, have diverse functions in different pathways involving angiogenesis, inflammation and apoptosis. The functional diversity is further illustrated in the association with various diseases through genetic mutations that do not affect aminoacylation or protein synthesis. Here we review the accumulated knowledge on how human tRNA synthetases used structural inventions to achieve functional expansions. PMID:19932696

  14. Evidence for two immunologically distinct acetyl-coenzyme A synthetases in yeast

    NASA Technical Reports Server (NTRS)

    Satyanarayana, T.; Mandel, A. D.; Klein, H. P.

    1974-01-01

    Evidence is presented that clearly establishes the presence of two acetyl-CoA synthetases in Saccharomyces cerevisiae, one elaborated under 'aerobic' conditions, the other under 'nonaerobic' conditions. The antibody produced by each enzyme is immunologically specific.

  15. Symmetry Groups for Linear Programming Relaxations of Orthogonal Array Problems

    DTIC Science & Technology

    2015-03-26

    Symmetry Groups for Linear Programming Relaxations of Orthogonal Array Problems THESIS MARCH 2015 David M. Arquette, Second Lieutenant, USAF AFIT-ENC...work of the U.S. Government and is not subject to copyright protection in the United States. AFIT-ENC-MS-15-M-003 SYMMETRY GROUPS FOR LINEAR...PUBLIC RELEASE; DISTRIBUTION UNLIMITED. AFIT-ENC-MS-15-M-003 SYMMETRY GROUPS FOR LINEAR PROGRAMMING RELAXATIONS OF ORTHOGONAL ARRAY PROBLEMS David M

  16. A few thoughts on proper orthogonal decomposition in turbulence

    NASA Astrophysics Data System (ADS)

    Podvin, Bérengère; Fraigneau, Yann

    2017-02-01

    Proper orthogonal decomposition was originally introduced in turbulence to identify large-scale patterns in turbulent flows. Over the years, several extensions have been formulated in order to strengthen its model-predictive abilities, with limited success in the case of fully developed turbulence. We argue that physics-based insight obtained from the proper orthogonal decomposition structures and other turbulence analysis techniques could lead to significant developments in that respect. Numerical results from channel flow simulations are used to illustrate our conjectures.

  17. Local unitary equivalence of quantum states and simultaneous orthogonal equivalence

    NASA Astrophysics Data System (ADS)

    Jing, Naihuan; Yang, Min; Zhao, Hui

    2016-06-01

    The correspondence between local unitary equivalence of bipartite quantum states and simultaneous orthogonal equivalence is thoroughly investigated and strengthened. It is proved that local unitary equivalence can be studied through simultaneous similarity under projective orthogonal transformations, and four parametrization independent algorithms are proposed to judge when two density matrices on ℂd1 ⊗ ℂd2 are locally unitary equivalent in connection with trace identities, Kronecker pencils, Albert determinants and Smith normal forms.

  18. Computerized tomography using a modified orthogonal tangent correction algorithm.

    PubMed

    Hsia, T C; Smith, S C; Lantz, B M

    1976-10-01

    A modified orthogonal tangent correction algorithm is presented for computerized tomography. The algorithm uses four X-rays scans spaced 45 degrees apart, to reconstruct a transverse axial image. The reconstruction procedure is interative in which image matrix elements are corrected by alternately matching the two sets of orthogonal scan data. The algorithm has been applied to phantom data as well as to video recorded fluoroscopic data.

  19. Expansion and orthogonalization of measured modes for structure identification

    NASA Technical Reports Server (NTRS)

    Smith, Suzanne Weaver

    1989-01-01

    The purpose was to investigate a new simultaneous expansion/orthogonalization method in comparison with two previously published expansion methods and a widely used orthogonalization technique. Each expansion method uses data from an analytical model of the structure to complete the estimate of the mode shape vectors. Berman and Nagy used Guyan expansion in their work with improving analytical models. In this method, modes are expanded one at a time, producing a set not orthogonal with respect to the mass matrix. Baruch and Bar Itzhack's optimal orthogonalization procedure was used to subsequently adjust the expanded modes. A second expansion technique was presented by O'Callahan, Avitabile, and Reimer and separately by Kammer. Again, modes are expanded individually and orthogonalized after expansion with the same optimal technique as above. Finally, a simultaneous expansion/orthogonalization method was developed from the orthogonal Procrustes problem of computational mathematics. In this method modes are optimally expanded as a set and orthogonal with respect to the mass matrix as a result. Two demonstation problems were selected for the comparison of the methods described. The first problem is an 8 degree of freedom spring-mass problem first presented by Kabe. Several conditions were examined for expansion method including the presence of errors in the measured data and in the analysis models. As a second demonstration problem, data from tests of laboratory scale model truss structures was expanded for system identification. Tests with a complete structure produced a correlated analysis model and the stiffness and mass matrices. Tests of various damaged configurations produced measured data for 6 modes at 14 dof locations.

  20. ONMCGP: Orthogonal Neighbourhood Mutation Cartesian Genetic Programming for Evolvable Hardware

    NASA Astrophysics Data System (ADS)

    I, Fuchuan N.; I, Yuanxiang L.; E, Peng K.

    2014-03-01

    Evolvable Hardware is facing the problems of scalability and stalling effect. This paper proposed a novel Orthogonal Neighbourhood Mutation (ONM) operator in Cartesian genetic programming (CGP), to reduce the stalling effect in CGP and improve the efficiency of the algorithms.The method incorporates with Differential Evolution strategy. Demonstrated by experiments on benchmark, the proposed Orthogonal Neighbourhood Search can jump out of Local optima, reduce the stalling effect in CGP and the algorithm convergence faster.

  1. Diffuse glutamine synthetase overexpression restricted to areas of peliosis in a β-catenin-activated hepatocellular adenoma: a potential pitfall in glutamine synthetase interpretation.

    PubMed

    Berry, Ryan S; Gullapalli, Rama R; Wu, Jin; Morris, Katherine; Hanson, Joshua A

    2014-08-01

    Hepatocellular adenomas have recently been classified into four subtypes based on molecular findings: hepatocyte nuclear factor 1α (HNF1α) inactivated, inflammatory/telangiectatic, β-catenin activated, and unclassifiable. β-catenin-activated adenomas have the potential for malignant transformation and are thus important to recognize. Diffuse glutamine synthetase immunohistochemical positivity has been shown to be a reliable surrogate marker for β-catenin activation, though variations in staining patterns may be difficult to interpret. We report a case of a peliotic adenoma that was morphologically consistent with a β-catenin wild-type hepatocellular adenoma but harbored a β-catenin mutation by molecular analysis. The tumor lacked nuclear β-catenin positivity and demonstrated a hitherto undescribed pattern of glutamine synthetase overexpression restricted to areas of peliosis with mostly negative staining in non-peliotic areas. This pattern was initially interpreted as physiologic and may represent a potential pitfall in glutamine synthetase interpretation.

  2. Lack of protective effect of thromboxane synthetase inhibitor (CGS-13080) on single dose radiated canine intestine.

    PubMed

    Barter, J F; Marlow, D; Kamath, R K; Harbert, J; Torrisi, J R; Barnes, W A; Potkul, R K; Newsome, J T; Delgado, G

    1991-03-01

    The effect of a thromboxane A2 synthetase inhibitor (CGS-13080) on canine intestine was studied using a single dose of radiation, and radioactive microspheres were used to determine resultant blood flow. Thromboxane A2 causes vasospasm and platelet aggregation and may play a dominant role in radiation injury. However, there was no effect on the intestinal blood flow diminution occurring after radiation in this laboratory model using this thromboxane A2 synthetase inhibitor.

  3. Lack of protective effect of thromboxane synthetase inhibitor (CGS-13080) on single dose radiated canine intestine

    SciTech Connect

    Barter, J.F.; Marlow, D.; Kamath, R.K.; Harbert, J.; Torrisi, J.R.; Barnes, W.A.; Potkul, R.K.; Newsome, J.T.; Delgado, G. )

    1991-03-01

    The effect of a thromboxane A2 synthetase inhibitor (CGS-13080) on canine intestine was studied using a single dose of radiation, and radioactive microspheres were used to determine resultant blood flow. Thromboxane A2 causes vasospasm and platelet aggregation and may play a dominant role in radiation injury. However, there was no effect on the intestinal blood flow diminution occurring after radiation in this laboratory model using this thromboxane A2 synthetase inhibitor.

  4. The identification of new cytosolic glutamine synthetase and asparagine synthetase genes in barley (Hordeum vulgare L.), and their expression during leaf senescence.

    PubMed

    Avila-Ospina, Liliana; Marmagne, Anne; Talbotec, Joël; Krupinska, Karin; Masclaux-Daubresse, Céline

    2015-04-01

    Glutamine synthetase and asparagine synthetase are two master enzymes involved in ammonium assimilation in plants. Their roles in nitrogen remobilization and nitrogen use efficiency have been proposed. In this report, the genes coding for the cytosolic glutamine synthetases (HvGS1) and asparagine synthetases (HvASN) in barley were identified. In addition to the three HvGS1 and two HvASN sequences previously reported, two prokaryotic-like HvGS1 and three HvASN cDNA sequences were identified. Gene structures were then characterized, obtaining full genomic sequences. The response of the five HvGS1 and five HvASN genes to leaf senescence was then studied. Developmental senescence was studied using primary and flag leaves. Dark-exposure or low-nitrate conditions were also used to trigger stress-induced senescence. Well-known senescence markers such as the chlorophyll and Rubisco contents were monitored in order to characterize senescence levels in the different leaves. The three eukaryotic-like HvGS1_1, HvGS1_2, and HvGS1_3 sequences showed the typical senescence-induced reduction in gene expression described in many plant species. By contrast, the two prokaryotic-like HvGS1_4 and HvGS1_5 sequences were repressed by leaf senescence, similar to the HvGS2 gene, which encodes the chloroplast glutamine synthetase isoenzyme. There was a greater contrast in the responses of the five HvASN and this suggested that these genes are needed for N remobilization in senescing leaves only when plants are well fertilized with nitrate. Responses of the HvASN sequences to dark-induced senescence showed that there are two categories of asparagine synthetases, one induced in the dark and the other repressed by the same conditions.

  5. Recurrent seizures and brain pathology after inhibition of glutamine synthetase in the hippocampus in rats.

    PubMed

    Eid, Tore; Ghosh, Arko; Wang, Yue; Beckström, Henning; Zaveri, Hitten P; Lee, Tih-Shih W; Lai, James C K; Malthankar-Phatak, Gauri H; de Lanerolle, Nihal C

    2008-08-01

    An excess of extracellular glutamate in the hippocampus has been linked to the generation of recurrent seizures and brain pathology in patients with medically intractable mesial temporal lobe epilepsy (MTLE). However, the mechanism which results in glutamate excess in MTLE remains unknown. We recently reported that the glutamate-metabolizing enzyme glutamine synthetase is deficient in the hippocampus in patients with MTLE, and we postulated that this deficiency is critically involved in the pathophysiology of the disease. To further explore the role of glutamine synthetase in MTLE we created a novel animal model of hippocampal glutamine synthetase deficiency by continuous (approximately 28 days) microinfusion of methionine sulfoximine (MSO: 0.625 to 2.5 microg/h) unilaterally into the hippocampus in rats. This treatment led to a deficiency in hippocampal glutamine synthetase activity by 82-97% versus saline. The majority (>95%) of the MSO-treated animals exhibited recurrent seizures that continued for several weeks. Some of the MSO-treated animals exhibited neuropathological features that were similar to mesial temporal sclerosis, such as hippocampal atrophy and patterned loss of hippocampal neurons. However, many MSO-treated animals displayed only minimal injury to the hippocampus, with no clear evidence of mesial temporal sclerosis. These findings support the hypothesis that a deficiency in hippocampal glutamine synthetase causes recurrent seizures, even in the absence of classical mesial temporal sclerosis, and that restoration of glutamine synthetase may represent a novel approach to therapeutic intervention in this disease.

  6. CTP synthetase and its role in phospholipid synthesis in the yeast Saccharomyces cerevisiae

    PubMed Central

    Chang, Yu-Fang; Carman, George M.

    2008-01-01

    CTP synthetase is a cytosolic-associated glutamine amidotransferase enzyme that catalyzes the ATP-dependent transfer of the amide nitrogen from glutamine to the C-4 position of UTP to form CTP. In the yeast Saccharomyces cerevisiae, the reaction product CTP is an essential precursor of all membrane phospholipids that are synthesized via the Kennedy (CDP-choline and CDP-ethanolamine branches) and CDP-diacylglycerol pathways. The URA7 and URA8 genes encode CTP synthetase in S. cerevisiae, and the URA7 gene is responsible for the majority of CTP synthesized in vivo. The CTP synthetase enzymes are allosterically regulated by CTP product inhibition. Mutations that alleviate this regulation result in an elevated cellular level of CTP and an increase in phospholipid synthesis via the Kennedy pathway. The URA7-encoded enzyme is phosphorylated by protein kinases A and C, and these phosphorylations stimulate CTP synthetase activity and increase cellular CTP levels and the utilization of the Kennedy pathway. The CTPS1 and CTPS2 genes that encode human CTP synthetase enzymes are functionally expressed in S. cerevisiae, and rescue the lethal phenotype of the ura7Δ ura8Δ double mutant that lacks CTP synthetase activity. The expression in yeast has revealed that the human CTPS1-encoded enzyme is also phosphorylated and regulated by protein kinases A and C. PMID:18439916

  7. Gain-Of-Function Mutational Activation of Human TRNA Synthetase Procytokine

    SciTech Connect

    Yang, X.L.; Kapoor, M.; Otero, F.J.; Slike, B.M.; Tsuruta, H.; Frausto, R.; Bates, A.; Ewalt, K.L.; Cheresh, D.A.; Schimmel, P.; /Scripps Res. Inst. /SLAC, SSRL

    2009-04-30

    Disease-causing mutations occur in genes for aminoacyl tRNA synthetases. That some mutations are dominant suggests a gain of function. Native tRNA synthetases, such as tyrosyl-tRNA synthetase (TyrRS) and tryptophanyl-tRNA synthetase, catalyze aminoacylation and are also procytokines that are activated by natural fragmentation. In principle, however, gain-of-function phenotypes could arise from mutational activation of synthetase procytokines. From crystal structure analysis, we hypothesized that a steric block of a critical Glu-Leu-Arg (ELR) motif in full-length TyrRS suppresses the cytokine activity of a natural fragment. To test this hypothesis, we attempted to uncover ELR in the procytokine by mutating a conserved tyrosine (Y341) that tethers ELR. Site-specific proteolytic cleavage and small-angle X-ray scattering established subtle opening of the structure by the mutation. Strikingly, four different assays demonstrated mutational activation of cytokine functions. The results prove the possibilities for constitutive gain-of-function mutations in tRNA synthetases.

  8. On the equivalence of dynamically orthogonal and bi-orthogonal methods: Theory and numerical simulations

    SciTech Connect

    Choi, Minseok; Sapsis, Themistoklis P.; Karniadakis, George Em

    2014-08-01

    The Karhunen–Lòeve (KL) decomposition provides a low-dimensional representation for random fields as it is optimal in the mean square sense. Although for many stochastic systems of practical interest, described by stochastic partial differential equations (SPDEs), solutions possess this low-dimensional character, they also have a strongly time-dependent form and to this end a fixed-in-time basis may not describe the solution in an efficient way. Motivated by this limitation of standard KL expansion, Sapsis and Lermusiaux (2009) [26] developed the dynamically orthogonal (DO) field equations which allow for the simultaneous evolution of both the spatial basis where uncertainty ‘lives’ but also the stochastic characteristics of uncertainty. Recently, Cheng et al. (2013) [28] introduced an alternative approach, the bi-orthogonal (BO) method, which performs the exact same tasks, i.e. it evolves the spatial basis and the stochastic characteristics of uncertainty. In the current work we examine the relation of the two approaches and we prove theoretically and illustrate numerically their equivalence, in the sense that one method is an exact reformulation of the other. We show this by deriving a linear and invertible transformation matrix described by a matrix differential equation that connects the BO and the DO solutions. We also examine a pathology of the BO equations that occurs when two eigenvalues of the solution cross, resulting in an instantaneous, infinite-speed, internal rotation of the computed spatial basis. We demonstrate that despite the instantaneous duration of the singularity this has important implications on the numerical performance of the BO approach. On the other hand, it is observed that the BO is more stable in nonlinear problems involving a relatively large number of modes. Several examples, linear and nonlinear, are presented to illustrate the DO and BO methods as well as their equivalence.

  9. Pivotal role of glutamine synthetase in ammonia detoxification.

    PubMed

    Hakvoort, Theodorus B M; He, Youji; Kulik, Wim; Vermeulen, Jacqueline L M; Duijst, Suzanne; Ruijter, Jan M; Runge, Jurgen H; Deutz, Nicolaas E P; Koehler, S Eleonore; Lamers, Wouter H

    2017-01-01

    Glutamine synthetase (GS) catalyzes condensation of ammonia with glutamate to glutamine. Glutamine serves, with alanine, as a major nontoxic interorgan ammonia carrier. Elimination of hepatic GS expression in mice causes only mild hyperammonemia and hypoglutaminemia but a pronounced decrease in the whole-body muscle-to-fat ratio with increased myostatin expression in muscle. Using GS-knockout/liver and control mice and stepwise increments of enterally infused ammonia, we show that ∼35% of this ammonia is detoxified by hepatic GS and ∼35% by urea-cycle enzymes, while ∼30% is not cleared by the liver, independent of portal ammonia concentrations ≤2 mmol/L. Using both genetic (GS-knockout/liver and GS-knockout/muscle) and pharmacological (methionine sulfoximine and dexamethasone) approaches to modulate GS activity, we further show that detoxification of stepwise increments of intravenously (jugular vein) infused ammonia is almost totally dependent on GS activity. Maximal ammonia-detoxifying capacity through either the enteral or the intravenous route is ∼160 μmol/hour in control mice. Using stable isotopes, we show that disposal of glutamine-bound ammonia to urea (through mitochondrial glutaminase and carbamoylphosphate synthetase) depends on the rate of glutamine synthesis and increases from ∼7% in methionine sulfoximine-treated mice to ∼500% in dexamethasone-treated mice (control mice, 100%), without difference in total urea synthesis. Hepatic GS contributes to both enteral and systemic ammonia detoxification. Glutamine synthesis in the periphery (including that in pericentral hepatocytes) and glutamine catabolism in (periportal) hepatocytes represents the high-affinity ammonia-detoxifying system of the body. The dependence of glutamine-bound ammonia disposal to urea on the rate of glutamine synthesis suggests that enhancing peripheral glutamine synthesis is a promising strategy to treat hyperammonemia. Because total urea synthesis does not depend on

  10. Spectrophotometric studies of acyl-coenzyme A synthetases of rat liver mitochondria

    PubMed Central

    Garland, P. B.; Yates, D. W.; Haddock, B. A.

    1970-01-01

    1. Deca-2,4,6,8-tetraenoic acid is a substrate for both ATP-specific (EC 6.2.1.2 or 3) and GTP-specific (EC 6.2.1.–) acyl-CoA synthetases of rat liver mitochondria. The enzymic synthesis of decatetraenoyl-CoA results in new spectral characteristics. The difference spectrum for the acyl-CoA minus free acid has a maximum at 376nm with εmM 34. Isosbestic points are at 345nm and 440nm. 2. The acylation of CoA by decatetraenoate in mitochondrial suspensions can be continuously measured with a dual-wavelength spectrophotometer. 3. By using this technique, three distinct types of acyl-CoA synthetase activity were demonstrated in rat liver mitochondria. One of these utilized added CoA and ATP, required added Mg2+ and corresponded to a previously described `external' acyl-CoA synthetase. The other two acyl-CoA synthetase activities utilized intramitochondrial CoA and did not require added Mg2+. Of these two `internal' acyl-CoA synthetases, one was insensitive to uncoupling agents, was inhibited by phosphate or arsenate, and corresponded to the GTP-specific enzyme. The other corresponded to the ATP-specific enzyme. 4. Atractylate inhibited the activity of the two internal acyl-CoA synthetases only when the energy source was added ATP. 5. The amount of intramitochondrial CoA acylated by decatetraenoate was independent of whether the internal ATP-specific or GTP-specific acyl-CoA synthetase was active. It is concluded that these two internal acyl-CoA synthetases have access to the same intramitochondrial pool of CoA. 6. The amount of intramitochondrial CoA that could be acylated with decatetraenoate was decreased by the addition of palmitoyl-dl-carnitine, 2-oxoglutarate, or pyruvate. These observations indicated that pyruvate dehydrogenase (EC 1.2.4.1), oxoglutarate dehydrogenase (EC 1.2.4.2), carnitine palmitoyltransferase (EC 2.3.1.–), citrate synthase (EC 4.1.3.7), and succinyl-CoA synthetase (EC 6.2.1.4) all have access to the same intramitochondrial pool of CoA as do

  11. Actinobacterial Acyl Coenzyme A Synthetases Involved in Steroid Side-Chain Catabolism

    PubMed Central

    Casabon, Israël; Swain, Kendra; Crowe, Adam M.

    2014-01-01

    Bacterial steroid catabolism is an important component of the global carbon cycle and has applications in drug synthesis. Pathways for this catabolism involve multiple acyl coenzyme A (CoA) synthetases, which activate alkanoate substituents for β-oxidation. The functions of these synthetases are poorly understood. We enzymatically characterized four distinct acyl-CoA synthetases from the cholate catabolic pathway of Rhodococcus jostii RHA1 and the cholesterol catabolic pathway of Mycobacterium tuberculosis. Phylogenetic analysis of 70 acyl-CoA synthetases predicted to be involved in steroid metabolism revealed that the characterized synthetases each represent an orthologous class with a distinct function in steroid side-chain degradation. The synthetases were specific for the length of alkanoate substituent. FadD19 from M. tuberculosis H37Rv (FadD19Mtb) transformed 3-oxo-4-cholesten-26-oate (kcat/Km = 0.33 × 105 ± 0.03 × 105 M−1 s−1) and represents orthologs that activate the C8 side chain of cholesterol. Both CasGRHA1 and FadD17Mtb are steroid-24-oyl-CoA synthetases. CasG and its orthologs activate the C5 side chain of cholate, while FadD17 and its orthologs appear to activate the C5 side chain of one or more cholesterol metabolites. CasIRHA1 is a steroid-22-oyl-CoA synthetase, representing orthologs that activate metabolites with a C3 side chain, which accumulate during cholate catabolism. CasI had similar apparent specificities for substrates with intact or extensively degraded steroid nuclei, exemplified by 3-oxo-23,24-bisnorchol-4-en-22-oate and 1β(2′-propanoate)-3aα-H-4α(3″-propanoate)-7aβ-methylhexahydro-5-indanone (kcat/Km = 2.4 × 105 ± 0.1 × 105 M−1 s−1 and 3.2 × 105 ± 0.3 × 105 M−1 s−1, respectively). Acyl-CoA synthetase classes involved in cholate catabolism were found in both Actinobacteria and Proteobacteria. Overall, this study provides insight into the physiological roles of acyl-CoA synthetases in steroid catabolism and

  12. Synthesis and site-directed fluorescence labeling of azido proteins using eukaryotic cell-free orthogonal translation systems.

    PubMed

    Quast, Robert B; Claussnitzer, Iris; Merk, Helmut; Kubick, Stefan; Gerrits, Michael

    2014-04-15

    Eukaryotic cell-free systems based on wheat germ and Spodoptera frugiperda insect cells were equipped with an orthogonal amber suppressor tRNA-synthetase pair to synthesize proteins with a site-specifically incorporated p-azido-l-phenylalanine residue in order to provide their chemoselective fluorescence labeling with azide-reactive dyes by Staudinger ligation. The specificity of incorporation and bioorthogonality of labeling within complex reaction mixtures was shown by means of translation and fluorescence detection of two model proteins: β-glucuronidase and erythropoietin. The latter contained the azido amino acid in proximity to a signal peptide for membrane translocation into endogenous microsomal vesicles of the insect cell-based system. The results indicate a stoichiometric incorporation of the azido amino acid at the desired position within the proteins. Moreover, the compatibility of cotranslational protein translocation, including glycosylation and amber suppression-based incorporation of p-azido-l-phenylalanine within a cell-free system, is demonstrated. The presented approach should be particularly useful for providing eukaryotic and membrane-associated proteins for investigation by fluorescence-based techniques.

  13. Expression of glutamine synthetase in the mouse kidney: localization in multiple epithelial cell types and differential regulation by hypokalemia.

    PubMed

    Verlander, Jill W; Chu, Diana; Lee, Hyun-Wook; Handlogten, Mary E; Weiner, I David

    2013-09-01

    Renal glutamine synthetase catalyzes the reaction of NH4+ with glutamate, forming glutamine and decreasing the ammonia available for net acid excretion. The purpose of the present study was to determine glutamine synthetase's specific cellular expression in the mouse kidney and its regulation by hypokalemia, a common cause of altered renal ammonia metabolism. Glutamine synthetase mRNA and protein were present in the renal cortex and in both the outer and inner stripes of the outer medulla. Immunohistochemistry showed glutamine synthetase expression throughout the entire proximal tubule and in nonproximal tubule cells. Double immunolabel with cell-specific markers demonstrated glutamine synthetase expression in type A intercalated cells, non-A, non-B intercalated cells, and distal convoluted tubule cells, but not in principal cells, type B intercalated cells, or connecting segment cells. Hypokalemia induced by feeding a nominally K+ -free diet for 12 days decreased glutamine synthetase expression throughout the entire proximal tubule and in the distal convoluted tubule and simultaneously increased glutamine synthetase expression in type A intercalated cells in both the cortical and outer medullary collecting duct. We conclude that glutamine synthetase is widely and specifically expressed in renal epithelial cells and that the regulation of expression differs in specific cell populations. Glutamine synthetase is likely to mediate an important role in renal ammonia metabolism.

  14. Homology modeling and molecular docking studies of Bacillomycin and Iturin synthetases with novel ligands for the production of therapeutic lipopeptides.

    PubMed

    Eswari, Jujjavarapu Satya; Dhagat, Swasti; Kaser, Shubham; Tiwari, Anoop

    2017-08-15

    Lipopeptide synthetases play an important role in the production of lipopeptides. Lipopeptides are molecules made up of peptides and fatty acid moieties and have shown to have a broad range of antimicrobial activity. As infectious diseases have caused severe health problems mainly resulting from the development of antibiotic resistant strains of disease causing microorganisms there is a need of alternatives to antibiotics. The lipopeptide synthetase of the corresponding lipopeptides can be used as templates to design these as drugs using computational techniques. The objective of this study was homology modeling and molecular docking of two lipopeptide synthetases, bacillomycin D synthetase and iturin A synthetase, with their ligands as a means of drug design. Schrödinger software was used for homology modeling and molecular docking. After the identification of ligands, molecular docking of these ligands with the lipopeptide (bacillomycin and iturin) synthetases was performed. The docking was tested on the parameters of docking score and glide energy. 5 out of 21 ligands were found to dock with bacillomycin D synthetase whereas 8 out of 20 ligands docked with the iturin A synthetase. The knowledge of the docking sites and docking characteristics of the lipopeptide synthetases mentioned in the paper with the ligands can provide advantages of high speed and reliability, reduced costs on chemicals and experiments and the ethical issues concerned with the use of animal models for screening of drug toxicity. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  15. Expression of glutamine synthetase in the mouse kidney: localization in multiple epithelial cell types and differential regulation by hypokalemia

    PubMed Central

    Chu, Diana; Lee, Hyun-Wook; Handlogten, Mary E.; Weiner, I. David

    2013-01-01

    Renal glutamine synthetase catalyzes the reaction of NH4+ with glutamate, forming glutamine and decreasing the ammonia available for net acid excretion. The purpose of the present study was to determine glutamine synthetase's specific cellular expression in the mouse kidney and its regulation by hypokalemia, a common cause of altered renal ammonia metabolism. Glutamine synthetase mRNA and protein were present in the renal cortex and in both the outer and inner stripes of the outer medulla. Immunohistochemistry showed glutamine synthetase expression throughout the entire proximal tubule and in nonproximal tubule cells. Double immunolabel with cell-specific markers demonstrated glutamine synthetase expression in type A intercalated cells, non-A, non-B intercalated cells, and distal convoluted tubule cells, but not in principal cells, type B intercalated cells, or connecting segment cells. Hypokalemia induced by feeding a nominally K+-free diet for 12 days decreased glutamine synthetase expression throughout the entire proximal tubule and in the distal convoluted tubule and simultaneously increased glutamine synthetase expression in type A intercalated cells in both the cortical and outer medullary collecting duct. We conclude that glutamine synthetase is widely and specifically expressed in renal epithelial cells and that the regulation of expression differs in specific cell populations. Glutamine synthetase is likely to mediate an important role in renal ammonia metabolism. PMID:23804452

  16. Differential inhibition of adenylylated and deadenylylated forms of M. tuberculosis glutamine synthetase as a drug discovery platform.

    PubMed

    Theron, A; Roth, R L; Hoppe, H; Parkinson, C; van der Westhuyzen, C W; Stoychev, S; Wiid, I; Pietersen, R D; Baker, B; Kenyon, C P

    2017-01-01

    Glutamine synthetase is a ubiquitous central enzyme in nitrogen metabolism that is controlled by up to four regulatory mechanisms, including adenylylation of some or all of the twelve subunits by adenylyl transferase. It is considered a potential therapeutic target for the treatment of tuberculosis, being essential for the growth of Mycobacterium tuberculosis, and is found extracellularly only in the pathogenic Mycobacterium strains. Human glutamine synthetase is not regulated by the adenylylation mechanism, so the adenylylated form of bacterial glutamine synthetase is of particular interest. Previously published reports show that, when M. tuberculosis glutamine synthetase is expressed in Escherichia coli, the E. coli adenylyl transferase does not optimally adenylylate the M. tuberculosis glutamine synthetase. Here, we demonstrate the production of soluble adenylylated M. tuberulosis glutamine synthetase in E. coli by the co-expression of M. tuberculosis glutamine synthetase and M. tuberculosis adenylyl transferase. The differential inhibition of adenylylated M. tuberulosis glutamine synthetase and deadenylylated M. tuberulosis glutamine synthetase by ATP based scaffold inhibitors are reported. Compounds selected on the basis of their enzyme inhibition were also shown to inhibit M. tuberculosis in the BACTEC 460TB™ assay as well as the intracellular inhibition of M. tuberculosis in a mouse bone-marrow derived macrophage assay.

  17. The gene encoding human glutathione synthetase (GSS) maps to the long arm of chromosome 20 at band 11.2

    SciTech Connect

    Webb, G.C.; Vaska, V.L.; Ford, J.H.

    1995-12-10

    Two forms of glutathione synthetase deficiency have been described. While one form is mild, causing hemolytic anemia, the other more severe form causes 5-oxoprolinuria with secondary neurological involvement. Despite the existence of two deficiency phenotypes, Southern blots hybridized with a glutathione synthetase cDNA suggest that there is a single glutathione synthetase gene in the human genome. Analysis of somatic cell hybrids showed the human glutathione synthetase gene (GSS) to be located on chromosome 20, and this assignment has been refined to subband 20q11.2 using in situ hybridization. 16 refs., 2 figs.

  18. Biochemical parameters of glutamine synthetase from Klebsiella aerogenes.

    PubMed Central

    Bender, R A; Janssen, K A; Resnick, A D; Blumenberg, M; Foor, F; Magasanik, B

    1977-01-01

    The glutamine synthetase (GS) from Klebsiella aerogenes is similar to that from Escherichia coli in several respects: (i) it is repressed by high levels of ammonia in the growth medium; (ii) its biosynthetic activity is greatly reduced by adenylylation; and (iii) adenylylation lowers the pH optimum and alters the response of the enzymes to various inhibitors in the gamma-glutamyl transferase (gammaGT) assay. There are, however, several important differences: (i) the isoactivity point for the adenylylated and non-adenylylated forms in the gammaGT assay occurs at pH 7.55 in K. aerogenes and at pH 7.15 in E. coli; (ii) the non-adenylylated form of the GS from K. aerogenes is stimulated by 60 mM MgCl2 in the gammaGT assay at pH 7.15. A biosynthetic reaction assay that correlates well with number of non-adenylylated enzyme subunits, as determined by the method of Mg2+ inhibition of the gammaGT assay, is described. Finally, we have found that it is necessary to use special methods to harvest growing cells to prevent changes in the adenylylation state of GS from occurring during harvesting. PMID:14104

  19. Insights into an Unusual Nonribosomal Peptide Synthetase Biosynthesis

    PubMed Central

    Binz, Tina M.; Maffioli, Sonia I.; Sosio, Margherita; Donadio, Stefano; Müller, Rolf

    2010-01-01

    The GE81112 tetrapeptides (1–3) represent a structurally unique class of antibiotics, acting as specific inhibitors of prokaryotic protein synthesis. Here we report the cloning and sequencing of the GE81112 biosynthetic gene cluster from Streptomyces sp. L-49973 and the development of a genetic manipulation system for Streptomyces sp. L-49973. The biosynthetic gene cluster for the tetrapeptide antibiotic GE81112 (getA-N) was identified within a 61.7-kb region comprising 29 open reading frames (open reading frames), 14 of which were assigned to the biosynthetic gene cluster. Sequence analysis revealed the GE81112 cluster to consist of six nonribosomal peptide synthetase (NRPS) genes encoding incomplete di-domain NRPS modules and a single free standing NRPS domain as well as genes encoding other biosynthetic and modifying proteins. The involvement of the cloned gene cluster in GE81112 biosynthesis was confirmed by inactivating the NRPS gene getE resulting in a GE81112 production abolished mutant. In addition, we characterized the NRPS A-domains from the pathway by expression in Escherichia coli and in vitro enzymatic assays. The previously unknown stereochemistry of most chiral centers in GE81112 was established from a combined chemical and biosynthetic approach. Taken together, these findings have allowed us to propose a rational model for GE81112 biosynthesis. The results further open the door to developing new derivatives of these promising antibiotic compounds by genetic engineering. PMID:20710026

  20. Novel acyl-CoA synthetase in adrenoleukodystrophy target tissues.

    PubMed

    Moriya-Sato, A; Hida, A; Inagawa-Ogashiwa, M; Wada, M R; Sugiyama, K; Shimizu, J; Yabuki, T; Seyama, Y; Hashimoto, N

    2000-12-09

    X-linked adrenoleukodystrophy (X-ALD) is a neurodegenerative disorder characterized by demyelination of white matter. The X-ALD gene product adrenoleukodystrophy protein (ALDP) is expressed broadly among various tissues. However, deficiency of functional ALDP exclusively impairs brain, adrenal gland, and testis. Thus, loss of ALDP function is assumed to involve inactivation of a putative mediating factor that functions in a tissue-specific manner. Here we cloned a mouse cDNA encoding a novel protein, Lipidosin, that possesses long-chain acyl-CoA synthetase (LCAS) activity. Lipidosin is expressed exclusively in mouse brain, adrenal gland, and testis, which are affected by X-ALD. LCAS activity of Lipidosin was diminished by mutation of conserved amino acids within the AMP-binding domain. Mutation of the Drosophila homologue of Lipidosin has been reported to cause neuronal degeneration. Thus, Lipidosin may mediate the link between ALDP dysfunction and the impairment of fatty acid metabolism in X-ALD. Copyright 2000 Academic Press.

  1. Nucleotide triphosphate promiscuity in Mycobacterium tuberculosis dethiobiotin synthetase.

    PubMed

    Salaemae, Wanisa; Yap, Min Y; Wegener, Kate L; Booker, Grant W; Wilce, Matthew C J; Polyak, Steven W

    2015-05-01

    Dethiobiotin synthetase (DTBS) plays a crucial role in biotin biosynthesis in microorganisms, fungi, and plants. Due to its importance in bacterial pathogenesis, and the absence of a human homologue, DTBS is a promising target for the development of new antibacterials desperately needed to combat antibiotic resistance. Here we report the first X-ray structure of DTBS from Mycobacterium tuberculosis (MtDTBS) bound to a nucleotide triphosphate (CTP). The nucleoside base is stabilized in its pocket through hydrogen-bonding interactions with the protein backbone, rather than amino acid side chains. This resulted in the unexpected finding that MtDTBS could utilise ATP, CTP, GTP, ITP, TTP, or UTP with similar Km and kcat values, although the enzyme had the highest affinity for CTP in competitive binding and surface plasmon resonance assays. This is in contrast to other DTBS homologues that preferentially bind ATP primarily through hydrogen-bonds between the purine base and the carboxamide side chain of a key asparagine. Mutational analysis performed alongside in silico experiments revealed a gate-keeper role for Asn175 in Escherichia coli DTBS that excludes binding of other nucleotide triphosphates. Here we provide evidence to show that MtDTBS has a broad nucleotide specificity due to the absence of the gate-keeper residue.

  2. Structural Biology of Non-Ribosomal Peptide Synthetases

    PubMed Central

    Miller, Bradley R.; Gulick, Andrew M.

    2016-01-01

    Summary The non-ribosomal peptide synthetases are modular enzymes that catalyze synthesis of important peptide products from a variety of standard and non-proteinogenic amino acid substrates. Within a single module are multiple catalytic domains that are responsible for incorporation of a single residue. After the amino acid is activated and covalently attached to an integrated carrier protein domain, the substrates and intermediates are delivered to neighboring catalytic domains for peptide bond formation or, in some modules, chemical modification. In the final module, the peptide is delivered to a terminal thioesterase domain that catalyzes release of the peptide product. This multi-domain modular architecture raises questions about the structural features that enable this assembly line synthesis in an efficient manner. The structures of the core component domains have been determined and demonstrate insights into the catalytic activity. More recently, multi-domain structures have been determined and are providing clues to the features of these enzyme systems that govern the functional interaction between multiple domains. This chapter describes the structures of NRPS proteins and the strategies that are being used to assist structural studies of these dynamic proteins, including careful consideration of domain boundaries for generation of truncated proteins and the use of mechanism-based inhibitors that trap interactions between the catalytic and carrier protein domains. PMID:26831698

  3. The prokaryotic FAD synthetase family: a potential drug target.

    PubMed

    Serrano, Ana; Ferreira, Patricia; Martínez-Júlvez, Marta; Medina, Milagros

    2013-01-01

    Disruption of cellular production of the flavin cofactors, flavin adenine mononucleotide (FMN) and flavin adenine dinucleotide(FAD) will prevent the assembly of a large number of flavoproteins and flavoenzymes involved in key metabolic processes in all types of organisms. The enzymes responsible for FMN and FAD production in prokaryotes and eukaryotes exhibit various structural characteristics to catalyze the same chemistry, a fact that converts the prokaryotic FAD synthetase (FADS) in a potential drug target for the development of inhibitors endowed with anti-pathogenic activity. The first step before searching for selective inhibitors of FADS is to understand the structural and functional mechanisms for the riboflavin kinase and FMN adenylyltransferase activities of the prokaryotic enzyme, and particularly to identify their differential functional characteristics with regard to the enzymes performing similar functions in other organisms, particularly humans. In this paper, an overview of the current knowledge of the structure-function relationships in prokaryotic FADS has been presented, as well as of the state of the art in the use of these enzymes as drug targets.

  4. In situ autoradiographic detection of folylpolyglutamate synthetase activity

    SciTech Connect

    Sussman, D.J.; Milman, G.; Osborne, C.; Shane, B.

    1986-11-01

    The enzyme folylpolyglutamate synthetase (FPGS) catalyzes the conversion of folate (pteroylmonoglutamate) to the polyglutamate forms (pteroylpolyglutamates) that are required for folate retention by mammalian cells. A rapid in situ autoradiographic assay for FPGS was developed which is based on the folate cofactor requirement of thymidylate synthase. Chinese hamster AUX B1 mutant cells lack FPGS activity and are unable to accumulate folate. As a result, the conversion of (6-/sup 3/H)deoxyuridine to thymidine via the thymidylate synthase reaction is impaired in AUX B1 cells and no detectable label is incorporated into DNA. In contrast, FPGS in wild-type Chinese hamster CHO cells causes folate retention and enables the incorporation of (6-/sup 3/H)deoxyuridine into DNA. Incorporation may be detected by autoradiography of monolayer cultures or of colonies replica plated onto polyester discs. Introduction of Escherichia coli FPGS into AUX B1 cells restores the activity of the thymidylate synthase pathway and demonstrates that the E. coli FPGS enzyme can provide pteroylpolyglutamates which functions in mammalian cells.

  5. Chitin synthetase in encysting Giardia lamblia and Entamoeba invadens

    SciTech Connect

    Das, S.; Gillin, F.D.

    1987-05-01

    Giardia lamblia (Gl) and Entamoeba invadens (Ei) are protozoan parasites with two morphologic stages in their life cycles. Motile trophozoites colonize the intestine of humans and reptiles respectively. Water resistant cysts, which can survive outside the host, transmit infection. In vitro cyst formation of Ei from trophozoites has been reported, and the authors have recently induced in vitro encystation of Gl. Although the cyst walls of both parasites contain chitin, it synthesis by encysting trophozoites has not been reported. The authors now show that encystation conditions greatly increase chitin synthetase (CS) specific activity (incorporation of /sup 3/H GlcNAc from UDP-GlcNAc into TCA-or alcohol-precipitable material). Extracts of encysting Gl incorporated 3.6 nmol/mg protein in 5 hr compared to < 0.005 in controls. Extracts of encysting Fi incorporated 4.8 n mol/mg protein, compared to 1.7 in the control. CS activity of both parasites requires preformed chitin. The Gl enzyme requires a reducing agent, is inhibited by digitonin and the CS inhibitors, polyoxin D and Nikkomycin, but not by tunicamycin. The product is digested by chitinase. Ei enzyme does not require a reducing agent and is stimulated by 1 mg/ml digitonin, but inhibited by higher concentrations. These studies demonstrate CS enzymes which may play important roles in encystation of Gl and Ei.

  6. The aminoacyl-tRNA synthetases of Drosophila melanogaster

    PubMed Central

    Lu, Jiongming; Marygold, Steven J; Gharib, Walid H; Suter, Beat

    2015-01-01

    Aminoacyl-tRNA synthetases (aaRSs) ligate amino acids to their cognate tRNAs, allowing them to decode the triplet code during translation. Through different mechanisms aaRSs also perform several non-canonical functions in transcription, translation, apoptosis, angiogenesis and inflammation. Drosophila has become a preferred system to model human diseases caused by mutations in aaRS genes, to dissect effects of reduced translation or non-canonical activities, and to study aminoacylation and translational fidelity. However, the lack of a systematic annotation of this gene family has hampered such studies. Here, we report the identification of the entire set of aaRS genes in the fly genome and we predict their roles based on experimental evidence and/or orthology. Further, we propose a new, systematic and logical nomenclature for aaRSs. We also review the research conducted on Drosophila aaRSs to date. Together, our work provides the foundation for further research in the fly aaRS field. PMID:26761199

  7. Cloning, expression, and purification of glutamine synthetase from Clostridum acetobutylicum

    SciTech Connect

    Usdin, K.P.; Zappe, H.; Jones, D.T.; Woods, D.R.

    1986-09-01

    A glutamine synthetase (GS) gene, glnA, from the gram-positive obligate anaerobe Clostridium acetobutylicum was cloned on recombinant plasmid pHZ200 and enabled Escherichia coli glnA deletion mutants to utilize (NH/sub 4/)/sub 2/ as a sole source of nitrogen. The cloned C. acetobutylicum gene was expressed from a regulatory region contained within the cloned DNA fragment. glnA expression was subject to nitrogen regulation in E. coli. This cloned glnA DNA did not enable an E. coli glnA ntrB ntrC deletion mutant to utilize arginine or low levels of glutamine as sole nitrogen sources, and failed to activate histidase activity in this strain which contained the Klebsiella aerogenes hut operon. The GS produced by pHZ200 was purified and had an apparent subunit molecular weight of approximately 59,000. There was no DNA or protein homology between the cloned C. acetobutylicum glnA gene and GS and the corresponding gene and GS from E. coli. The C. acetobutylicum GS was inhibited by Mg/sup 2 +/ in the ..gamma..-glutamyl transferase assay, but there was no evidence that the GS was adenylylated.

  8. Versatility of acyl-acyl carrier protein synthetases

    DOE PAGES

    Beld, Joris; Finzel, Kara; Burkart, Michael D.

    2014-10-09

    The acyl carrier protein (ACP) requires posttranslational modification with a 4'-phosphopantetheine arm for activity, and this thiol-terminated modification carries cargo between enzymes in ACP-dependent metabolic pathways. In this paper, we show that acyl-ACP synthetases (AasSs) from different organisms are able to load even, odd, and unnatural fatty acids onto E. coli ACP in vitro. Vibrio harveyi AasS not only shows promiscuity for the acid substrate, but also is active upon various alternate carrier proteins. AasS activity also extends to functional activation in living organisms. We show that exogenously supplied carboxylic acids are loaded onto ACP and extended by the E.more » coli fatty acid synthase, including unnatural fatty acid analogs. These analogs are further integrated into cellular lipids. Finally, in vitro characterization of four different adenylate-forming enzymes allowed us to disambiguate CoA-ligases and AasSs, and further in vivo studies show the potential for functional application in other organisms.« less

  9. The complex evolutionary history of aminoacyl-tRNA synthetases

    PubMed Central

    Chaliotis, Anargyros; Vlastaridis, Panayotis; Mossialos, Dimitris; Ibba, Michael; Becker, Hubert D.

    2017-01-01

    Abstract Aminoacyl-tRNA synthetases (AARSs) are a superfamily of enzymes responsible for the faithful translation of the genetic code and have lately become a prominent target for synthetic biologists. Our large-scale analysis of >2500 prokaryotic genomes reveals the complex evolutionary history of these enzymes and their paralogs, in which horizontal gene transfer played an important role. These results show that a widespread belief in the evolutionary stability of this superfamily is misconceived. Although AlaRS, GlyRS, LeuRS, IleRS, ValRS are the most stable members of the family, GluRS, LysRS and CysRS often have paralogs, whereas AsnRS, GlnRS, PylRS and SepRS are often absent from many genomes. In the course of this analysis, highly conserved protein motifs and domains within each of the AARS loci were identified and used to build a web-based computational tool for the genome-wide detection of AARS coding sequences. This is based on hidden Markov models (HMMs) and is available together with a cognate database that may be used for specific analyses. The bioinformatics tools that we have developed may also help to identify new antibiotic agents and targets using these essential enzymes. These tools also may help to identify organisms with alternative pathways that are involved in maintaining the fidelity of the genetic code. PMID:28180287

  10. Regulation of Acetyl Coenzyme A Synthetase in Escherichia coli

    PubMed Central

    Kumari, Suman; Beatty, Christine M.; Browning, Douglas F.; Busby, Stephen J. W.; Simel, Erica J.; Hovel-Miner, Galadriel; Wolfe, Alan J.

    2000-01-01

    Cells of Escherichia coli growing on sugars that result in catabolite repression or amino acids that feed into glycolysis undergo a metabolic switch associated with the production and utilization of acetate. As they divide exponentially, these cells excrete acetate via the phosphotransacetylase-acetate kinase pathway. As they begin the transition to stationary phase, they instead resorb acetate, activate it to acetyl coenzyme A (acetyl-CoA) by means of the enzyme acetyl-CoA synthetase (Acs) and utilize it to generate energy and biosynthetic components via the tricarboxylic acid cycle and the glyoxylate shunt, respectively. Here, we present evidence that this switch occurs primarily through the induction of acs and that the timing and magnitude of this induction depend, in part, on the direct action of the carbon regulator cyclic AMP receptor protein (CRP) and the oxygen regulator FNR. It also depends, probably indirectly, upon the glyoxylate shunt repressor IclR, its activator FadR, and many enzymes involved in acetate metabolism. On the basis of these results, we propose that cells induce acs, and thus their ability to assimilate acetate, in response to rising cyclic AMP levels, falling oxygen partial pressure, and the flux of carbon through acetate-associated pathways. PMID:10894724

  11. Small Alarmone Synthetases as novel bacterial RNA-binding proteins.

    PubMed

    Hauryliuk, Vasili; Atkinson, Gemma C

    2017-08-18

    The alarmone nucleotides guanosine pentaphosphate (pppGpp) and tetraphosphate (ppGpp), collectively referred to as (p)ppGpp, are key regulators of bacterial growth, stress adaptation, antibiotic tolerance and pathogenicity. We have recently shown that the Small Alarmone Synthetase (SAS) RelQ from the Gram-positive pathogen Enterococcus faecalis has an RNA-binding activity (Beljantseva et al. 2017). RelQ's activities as an enzyme and as a RNA-binding protein are mutually incompatible: binding of single-stranded RNA potently inhibits (p)ppGpp synthesis in a sequence-specific manner, and RelQ's enzymatic activity destabilizes the RNA:RelQ complex. RelQ's allosteric regulator, pppGpp, destabilizes RNA binding and activates RelQ's enzymatic activity. Since SAS enzymes are widely distributed in bacteria, and, as it has been discovered recently, are also mobilized by phages (Dedrick et al. 2017), RNA binding to SAS is could be a wide-spread mechanism. The initial discovery raises numerous questions regarding RNA-binding function of the SAS enzymes: What is the molecular mechanism underlying the incompatibility of RNA:SAS complex formation with pppGpp binding and (p)ppGpp synthesis? What are the RNA targets in living cells? What is the regulatory output of the system - (p)ppGpp synthesis, modulation of RNA structure and function, or both?

  12. Holocarboxylase synthetase: correlation of protein localisation with biological function.

    PubMed

    Bailey, L M; Wallace, J C; Polyak, S W

    2010-04-01

    Holocarboxylase synthetase (HCS) governs the cellular fate of the essential micronutrient biotin (Vitamin H or B7). HCS is responsible for attaching biotin onto the biotin-dependent enzymes that reside in the cytoplasm and mitochondria. Evidence for an alternative role, viz the regulation of gene expression, has also been reported. Recent immunohistochemical studies reported HCS is primarily nuclear, inconsistent with the location of HCS activity. Improved understanding of biotin biology demands greater knowledge about HCS. Here, we investigated the localisation of HCS and its isoforms. Three variants were observed that differ at the N-terminus. All HCS isoforms were predominantly non-nuclear, consistent with the distribution of biotin protein ligase activity. Unlike the longer constructs, the Met(58) isoform was also detected in the nucleus--a novel observation suggesting shuttling activity between nucleus and cytoplasm. We resolved that the previous controversies in the literature are due to specificity and detection limitations that arise when using partially purified antibodies. 2010. Published by Elsevier Inc. All rights reserved.

  13. Secondary NAD+ deficiency in the inherited defect of glutamine synthetase.

    PubMed

    Hu, Liyan; Ibrahim, Khalid; Stucki, Martin; Frapolli, Michele; Shahbeck, Noora; Chaudhry, Farrukh A; Görg, Boris; Häussinger, Dieter; Penberthy, W Todd; Ben-Omran, Tawfeg; Häberle, Johannes

    2015-11-01

    Glutamine synthetase (GS) deficiency is an ultra-rare inborn error of amino acid metabolism that has been described in only three patients so far. The disease is characterized by neonatal onset of severe encephalopathy, low levels of glutamine in blood and cerebrospinal fluid, chronic moderate hyperammonemia, and an overall poor prognosis in the absence of an effective treatment. Recently, enteral glutamine supplementation was shown to be a safe and effective therapy for this disease but there are no data available on the long-term effects of this intervention. The amino acid glutamine, severely lacking in this disorder, is central to many metabolic pathways in the human organism and is involved in the synthesis of nicotinamide adenine dinucleotide (NAD(+)) starting from tryptophan or niacin as nicotinate, but not nicotinamide. Using fibroblasts, leukocytes, and immortalized peripheral blood stem cells (PBSC) from a patient carrying a GLUL gene point mutation associated with impaired GS activity, we tested whether glutamine deficiency in this patient results in NAD(+) depletion and whether it can be rescued by supplementation with glutamine, nicotinamide or nicotinate. The present study shows that congenital GS deficiency is associated with NAD(+) depletion in fibroblasts, leukocytes and PBSC, which may contribute to the severe clinical phenotype of the disease. Furthermore, it shows that NAD(+) depletion can be rescued by nicotinamide supplementation in fibroblasts and leukocytes, which may open up potential therapeutic options for the treatment of this disorder.

  14. The plastidial folylpolyglutamate synthetase and root apical meristem maintenance

    PubMed Central

    Srivastava, Avinash C; Tang, Yuhong; Díaz de la Garza, Rocío I

    2011-01-01

    Folylpolyglutamate synthetase (FPGS) catalyzes the attachment of glutamate residues to the folate molecule in plants. Three isoforms of FPGS have been identified in Arabidopsis and these are localized in the plastid (AtDFB), mitochondria (AtDFC) and cytosol (AtDFD). We recently determined that mutants in the AtDFB (At5G05980) gene disrupt primary root development in Arabidopsis thaliana seedlings. Transient expression of AtDFB-green fluorescent protein (GFP) fusion under the control of the native AtDFB promoter in Nicotiana tabacum leaf epidermal cells verified the plastid localization of AtDFB. Furthermore, low concentrations of methotrexate (MTX), a compound commonly used as a folate antagonist in plant and mammalian cells induced primary root defects in wild type seedlings that were similar to atdfb. In addition, atdfb seedlings were more sensitive to MTX when compared to wild type. Quantitative (q) RT-PCR showed lower transcript levels of the mitochondrial and cytosolic FPGS in roots of 7-day-old atdfb seedling suggesting feedback regulation of AtDFB on the expression of other FPGS isoforms during early seedling development. The primary root defects of atdfb, which can be traced in part to altered quiescent center (QC) identity, pave the way for future studies that could link cell type specific folate and FPGS isoform requirements to whole organ development. PMID:21502816

  15. The enterococcal cytolysin synthetase has an unanticipated lipid kinase fold.

    PubMed

    Dong, Shi-Hui; Tang, Weixin; Lukk, Tiit; Yu, Yi; Nair, Satish K; van der Donk, Wilfred A

    2015-07-30

    The enterococcal cytolysin is a virulence factor consisting of two post-translationally modified peptides that synergistically kill human immune cells. Both peptides are made by CylM, a member of the LanM lanthipeptide synthetases. CylM catalyzes seven dehydrations of Ser and Thr residues and three cyclization reactions during the biosynthesis of the cytolysin large subunit. We present here the 2.2 Å resolution structure of CylM, the first structural information on a LanM. Unexpectedly, the structure reveals that the dehydratase domain of CylM resembles the catalytic core of eukaryotic lipid kinases, despite the absence of clear sequence homology. The kinase and phosphate elimination active sites that affect net dehydration are immediately adjacent to each other. Characterization of mutants provided insights into the mechanism of the dehydration process. The structure is also of interest because of the interactions of human homologs of lanthipeptide cyclases with kinases such as mammalian target of rapamycin.

  16. Application of joint orthogonal bases in compressive sensing ghost image

    NASA Astrophysics Data System (ADS)

    Fan, Xiang; Chen, Yi; Cheng, Zheng-dong; Liang, Zheng-yu; Zhu, Bin

    2016-11-01

    Sparse decomposition is one of the core issue of compressive sensing ghost image. At this stage, traditional methods still have the problems of poor sparsity and low reconstruction accuracy, such as discrete fourier transform and discrete cosine transform. In order to solve these problems, joint orthogonal bases transform is proposed to optimize ghost imaging. First, introduce the principle of compressive sensing ghost imaging and point out that sparsity is related to the minimum sample data required for imaging. Then, analyze the development and principle of joint orthogonal bases in detail and find out it can use less nonzero coefficients to reach the same identification effect as other methods. So, joint orthogonal bases transform is able to provide the sparsest representation. Finally, the experimental setup is built in order to verify simulation results. Experimental results indicate that the PSNR of joint orthogonal bases is much higher than traditional methods by using same sample data in compressive sensing ghost image.Therefore, joint orthogonal bases transform can realize better imaging quality under less sample data, which can satisfy the system requirements of convenience and rapid speed in ghost image.

  17. A modular strategy for engineering orthogonal chimeric RNA transcription regulators

    PubMed Central

    Takahashi, Melissa K.; Lucks, Julius B.

    2013-01-01

    Antisense RNA transcription attenuators are a key component of the synthetic biology toolbox, with their ability to serve as building blocks for both signal integration logic circuits and transcriptional cascades. However, a central challenge to building more sophisticated RNA genetic circuitry is creating larger families of orthogonal attenuators that function independently of each other. Here, we overcome this challenge by developing a modular strategy to create chimeric fusions between the engineered transcriptional attenuator from plasmid pT181 and natural antisense RNA translational regulators. Using in vivo gene expression assays in Escherichia coli, we demonstrate our ability to create chimeric attenuators by fusing sequences from five different translational regulators. Mutagenesis of these functional attenuators allowed us to create a total of 11 new chimeric attenutaors. A comprehensive orthogonality test of these culminated in a 7 × 7 matrix of mutually orthogonal regulators. A comparison between all chimeras tested led to design principles that will facilitate further engineering of orthogonal RNA transcription regulators, and may help elucidate general principles of non-coding RNA regulation. We anticipate that our strategy will accelerate the development of even larger families of orthogonal RNA transcription regulators, and thus create breakthroughs in our ability to construct increasingly sophisticated RNA genetic circuitry. PMID:23761434

  18. Positroid stratification of orthogonal Grassmannian and ABJM amplitudes

    NASA Astrophysics Data System (ADS)

    Kim, Joonho; Lee, Sangmin

    2014-09-01

    A novel understanding of scattering amplitudes in terms of on-shell diagrams and positive Grassmannian has been recently established for four dimensional Yang-Mills theories and three dimensional Chern-Simons theories of ABJM type. We give a detailed construction of the positroid stratification of orthogonal Grassmannian relevant for ABJM amplitudes. On-shell diagrams are classified by pairing of external particles. We introduce a combinatorial aid called `OG tableaux' and map each equivalence class of on-shell diagrams to a unique tableau. The on-shell diagrams related to each other through BCFW bridging are naturally grouped by the OG tableaux. Introducing suitably ordered BCFW bridges and positive coordinates, we construct the complete coordinate charts to cover the entire positive orthogonal Grassmannian for arbitrary number of external particles. The graded counting of OG tableaux suggests that the positive orthogonal Grassmannian constitutes a combinatorial polytope.

  19. Investigation of formation mechanisms of chips in orthogonal cutting process

    NASA Astrophysics Data System (ADS)

    Ma, W.

    2012-08-01

    This work investigates the formation mechanisms of chips in orthogonal cutting of mild steel and the transformation conditions between various morphology chips. It is supposed that the modeling material follows the Johnson-Cook constitutive model. In orthogonal cutting process, both the plastic flow and the instability behaviors of chip materials are caused by the plane strain loadings. Therefore, the general instability behaviors of materials in plane strain state are first analyzed with linear perturbation method and a universal instability criterion is established. Based on the analytical results, the formation mechanisms of chips and the transformation conditions between continuous and serrated chips are further studied by instability phase diagram method. The results show that the chip formation strongly depends on the intensity ratios between shear and normal stresses. The ratios of dissipative rates of plastic work done by compression and shear stresses govern the transformation from continuous to serrated chips. These results are verified by the numerical simulations on the orthogonal cutting process.

  20. Experimental quantum-cryptography scheme based on orthogonal states

    NASA Astrophysics Data System (ADS)

    Avella, Alessio; Brida, Giorgio; Degiovanni, Ivo Pietro; Genovese, Marco; Gramegna, Marco; Traina, Paolo

    2010-12-01

    Since, in general, nonorthogonal states cannot be cloned, any eavesdropping attempt in a quantum-communication scheme using nonorthogonal states as carriers of information introduces some errors in the transmission, leading to the possibility of detecting the spy. Usually, orthogonal states are not used in quantum-cryptography schemes since they can be faithfully cloned without altering the transmitted data. Nevertheless, L. Goldberg and L. Vaidman [Phys. Rev. Lett.PRLTAO0031-900710.1103/PhysRevLett.75.1239 75, 1239 (1995)] proposed a protocol in which, even if the data exchange is realized using two orthogonal states, any attempt to eavesdrop is detectable by the legal users. In this scheme the orthogonal states are superpositions of two localized wave packets traveling along separate channels. Here we present an experiment realizing this scheme.

  1. Experimental quantum-cryptography scheme based on orthogonal states

    SciTech Connect

    Avella, Alessio; Brida, Giorgio; Degiovanni, Ivo Pietro; Genovese, Marco; Gramegna, Marco; Traina, Paolo

    2010-12-15

    Since, in general, nonorthogonal states cannot be cloned, any eavesdropping attempt in a quantum-communication scheme using nonorthogonal states as carriers of information introduces some errors in the transmission, leading to the possibility of detecting the spy. Usually, orthogonal states are not used in quantum-cryptography schemes since they can be faithfully cloned without altering the transmitted data. Nevertheless, L. Goldberg and L. Vaidman [Phys. Rev. Lett. 75, 1239 (1995)] proposed a protocol in which, even if the data exchange is realized using two orthogonal states, any attempt to eavesdrop is detectable by the legal users. In this scheme the orthogonal states are superpositions of two localized wave packets traveling along separate channels. Here we present an experiment realizing this scheme.

  2. Bio-orthogonally Deciphered Binary Nanoemitters for Tumor Diagnostics.

    PubMed

    An, Hong-Wei; Qiao, Sheng-Lin; Li, Li-Li; Yang, Chao; Lin, Yao-Xin; Wang, Yi; Qiao, Zeng-Ying; Wang, Lei; Wang, Hao

    2016-08-03

    Bioinspired design concept has been recognized as one of the most promising strategies for discovering new biomaterials. However, smart biomaterials that are of growing interests in biomedical field need biological processability to meet their emergent applications in vivo. Herein, a new bio-orthogonally deciphered approach has been demonstrated for modulating optical properties of nanomaterials in living systems. The self-assembled nanoemitters based on cyanine-pyrene molecule 1 with inert optical property are designed and prepared. The structure and optical feature of the nanoemitters 1 can be efficiently and reliably modulated by a unique bio-orthogonal mechanism with abundant glutathione (GSH) as an activator. As a result, the self-assembled nanoemitters 1 spontaneously exhibits binary emissions for high-performance tumor imaging in vivo. We believe that this bio-orthogonally deciphered strategy opens a new avenue for designing variable smart biomaterials or devices in biomedical applications.

  3. All Possible Cayley-Klein Contractions of Quantum Orthogonal Groups

    SciTech Connect

    Gromov, N.A.; Kuratov, V.V.

    2005-10-01

    Spaces of constant curvature and their motion groups are described most naturally in the Cartesian basis. All these motion groups, also known as CK groups, are obtained from an orthogonal group by contractions and analytical continuations. On the other hand, quantum deformation of orthogonal group SO(N) is most easily performed in the so-called symplectic basis. We reformulate its standard quantum deformation to the Cartesian basis and obtain all possible contractions of quantum orthogonal group SO{sub q}(N) for both untouched and transformed deformation parameters. It turned out that, similar to the undeformed case, all CK contractions of SO{sub q}(N) are realized. An algorithm for obtaining nonequivalent (as Hopf algebra) contracted quantum groups is suggested. Contractions of SO{sub q}(N), N = 3, 4, 5, are regarded as examples.

  4. Simultaneous expansion and orthogonalization of measured modes for structure identification

    NASA Technical Reports Server (NTRS)

    Smith, Suzanne Weaver; Beattie, Christopher A.

    1990-01-01

    Tests of large structures on-orbit will be performed with measurements at a relatively few structure points. Values for the unmeasured degrees of freedom (dofs) can be estimated based on measured dofs and analytical model dynamic information. These 'expanded' mode shapes are useful for optimal-update identification and damage location as well as test/analysis correlation. A new method of expansion for test mode shape vectors is developed from the orthogonal Procrustes problem from computational linear algebra. A subspace defined by the set of measured dofs is compared to a subspace defined by mode shapes from an analytical model of the structure. The method simultaneously expands and orthogonalizes the mode shape vectors. Two demonstration problems are used to compare the new method to current expansion techniques. One demonstration uses test data from a laboratory scale-model truss structure. Performance of the new method is comparable or superior to that of the previous expansion methods which require separate orthogonalization.

  5. Semi-orthogonal wavelets for elliptic variational problems

    SciTech Connect

    Hardin, D.P.; Roach, D.W.

    1998-04-01

    In this paper the authors give a construction of wavelets which are (a) semi-orthogonal with respect to an arbitrary elliptic bilinear form a({center_dot},{center_dot}) on the Sobolev space H{sub 0}{sup 1}((0, L)) and (b) continuous and piecewise linear on an arbitrary partition of [0, L]. They illustrate this construction using a model problem. They also construct alpha-orthogonal Battle-Lemarie type wavelets which fully diagonalize the Galerkin discretized matrix for the model problem with domain IR. Finally they describe a hybrid basis consisting of a combination of elements from the semi-orthogonal wavelet basis and the hierarchical Schauder basis. Numerical experiments indicate that this basis leads to robust scalable Galerkin discretizations of the model problem which remain well-conditioned independent of {epsilon}, L, and the refinement level K.

  6. Locally indistinguishable orthogonal product bases in arbitrary bipartite quantum system

    PubMed Central

    Xu, Guang-Bao; Yang, Ying-Hui; Wen, Qiao-Yan; Qin, Su-Juan; Gao, Fei

    2016-01-01

    As we know, unextendible product basis (UPB) is an incomplete basis whose members cannot be perfectly distinguished by local operations and classical communication. However, very little is known about those incomplete and locally indistinguishable product bases that are not UPBs. In this paper, we first construct a series of orthogonal product bases that are completable but not locally distinguishable in a general m ⊗ n (m ≥ 3 and n ≥ 3) quantum system. In particular, we give so far the smallest number of locally indistinguishable states of a completable orthogonal product basis in arbitrary quantum systems. Furthermore, we construct a series of small and locally indistinguishable orthogonal product bases in m ⊗ n (m ≥ 3 and n ≥ 3). All the results lead to a better understanding of the structures of locally indistinguishable product bases in arbitrary bipartite quantum system. PMID:27503634

  7. Superposition of an orthogonal oscillation to study anisotropy in polymers

    NASA Astrophysics Data System (ADS)

    Coletti, Marco; Pepi, Renzo

    2014-05-01

    Rheology is routinely used to assess visco-elastic properties and structure of polymers, both in the melt state and in solution. Standard rheometers, though, can apply shear or oscillation in one direction, only. Several systems show a clear anisotropic behavior when tested along different directions, but a typical rheometer cannot perform such 2D deformation. In this paper we propose a different approach to such a characterization, where a controlled oscillation is applied in an orthogonal direction with respect to shear or oscillation. This technique is capable of revealing additional information on how samples behave in two directions, since it is either possible to superimpose the orthogonal oscillation to a constant shear (Orthogonal SuperPosition: OSP) or to another oscillation in the shear direction (2D SAOS).

  8. Modelling Trends in Ordered Correspondence Analysis Using Orthogonal Polynomials.

    PubMed

    Lombardo, Rosaria; Beh, Eric J; Kroonenberg, Pieter M

    2016-06-01

    The core of the paper consists of the treatment of two special decompositions for correspondence analysis of two-way ordered contingency tables: the bivariate moment decomposition and the hybrid decomposition, both using orthogonal polynomials rather than the commonly used singular vectors. To this end, we will detail and explain the basic characteristics of a particular set of orthogonal polynomials, called Emerson polynomials. It is shown that such polynomials, when used as bases for the row and/or column spaces, can enhance the interpretations via linear, quadratic and higher-order moments of the ordered categories. To aid such interpretations, we propose a new type of graphical display-the polynomial biplot.

  9. Generalized Discriminant Orthogonal Nonnegative Tensor Factorization for Facial Expression Recognition

    PubMed Central

    XiuJun, Zhang; Chang, Liu

    2014-01-01

    In order to overcome the limitation of traditional nonnegative factorization algorithms, the paper presents a generalized discriminant orthogonal non-negative tensor factorization algorithm. At first, the algorithm takes the orthogonal constraint into account to ensure the nonnegativity of the low-dimensional features. Furthermore, the discriminant constraint is imposed on low-dimensional weights to strengthen the discriminant capability of the low-dimensional features. The experiments on facial expression recognition have demonstrated that the algorithm is superior to other non-negative factorization algorithms. PMID:24982970

  10. An online incremental orthogonal component analysis method for dimensionality reduction.

    PubMed

    Zhu, Tao; Xu, Ye; Shen, Furao; Zhao, Jinxi

    2017-01-01

    In this paper, we introduce a fast linear dimensionality reduction method named incremental orthogonal component analysis (IOCA). IOCA is designed to automatically extract desired orthogonal components (OCs) in an online environment. The OCs and the low-dimensional representations of original data are obtained with only one pass through the entire dataset. Without solving matrix eigenproblem or matrix inversion problem, IOCA learns incrementally from continuous data stream with low computational cost. By proposing an adaptive threshold policy, IOCA is able to automatically determine the dimension of feature subspace. Meanwhile, the quality of the learned OCs is guaranteed. The analysis and experiments demonstrate that IOCA is simple, but efficient and effective.

  11. Empirical orthogonal functions and normal modes. [for atmospheric data studies

    NASA Technical Reports Server (NTRS)

    North, G. R.

    1984-01-01

    An attempt to provide physical insight into the empirical orthogonal function (EOF) representation of data fields by the study of fields generated by linear stochastic models is presented in this paper. In a large class of these models, the EOFs at individual Fourier frequencies coincide with the orthogonal mechanical modes of the system - provided they exist. The precise mathematical criteria for this coincidence are derived and a physical interpretation is provided. A scheme possibly useful in forecasting is formally constructed for representing any stochastic field by a linear Hermitian model forced by noise.

  12. Uniform Asymptotics of Orthogonal Polynomials Arising from Coherent States

    NASA Astrophysics Data System (ADS)

    Dai, Dan; Hu, Weiying; Wang, Xiang-Sheng

    2015-08-01

    In this paper, we study a family of orthogonal polynomials {φ_n(z)} arising from nonlinear coherent states in quantum optics. Based on the three-term recurrence relation only, we obtain a uniform asymptotic expansion of φ_n(z) as the polynomial degree n tends to infinity. Our asymptotic results suggest that the weight function associated with the polynomials has an unusual singularity, which has never appeared for orthogonal polynomials in the Askey scheme. Our main technique is the Wang and Wong's difference equation method. In addition, the limiting zero distribution of the polynomials φ_n(z) is provided.

  13. Connection preserving deformations and q-semi-classical orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Ormerod, Christopher M.; Witte, N. S.; Forrester, Peter J.

    2011-09-01

    We present a framework for the study of q-difference equations satisfied by q-semi-classical orthogonal systems. As an example, we identify the q-difference equation satisfied by a deformed version of the little q-Jacobi polynomials as a gauge transformation of a special case of the associated linear problem for q-PVI. We obtain a parametrization of the associated linear problem in terms of orthogonal polynomial variables and find the relation between this parametrization and that of Jimbo and Sakai.

  14. Post-functionalization of polymers via orthogonal ligation chemistry.

    PubMed

    Goldmann, Anja S; Glassner, Mathias; Inglis, Andrew J; Barner-Kowollik, Christopher

    2013-05-27

    The establishment of advanced living/controlled polymerization protocols allows for engineering synthetic polymers in a precise fashion. Combining advanced living/controlled polymerization techniques with highly efficient coupling chemistries facilitates quantitative, modular, and orthogonal functionalization of synthetic polymer strands at their chain termini as well as side-chain functionalization. The review highlights the current status of selected post-functionalization techniques of polymers via orthogonal ligation chemistries, major characteristics of the specific transformation chemistry, as well as the characterization of the products.

  15. Regulation of synthesis of glutamate dehydrogenase and glutamine synthetase in micro-organisms

    PubMed Central

    Pateman, J. A.

    1969-01-01

    1. Aspergillus nidulans, Neurospora crassa and Escherichia coli were grown on media containing a range of concentrations of nitrate, or ammonia, or urea, or l-glutamate, or l-glutamine as the sole source of nitrogen and the glutamate dehydrogenate and glutamine synthetase of the cells measured. 2. Aspergillus, Neurospora and Escherichia coli cells, grown on l-glutamate or on high concentrations of ammonia or on high concentrations of urea, possessed low glutamate dehydrogenase activity compared with cells grown on other nitrogen sources. 3. Aspergillus, Neurospora and Escherichia coli cells grown on l-glutamate possessed high glutamine synthetase activity compared with cells grown on other nitrogen sources. 4. The hypothesis is proposed that in Aspergillus, Neurospora and Escherichia coli l-glutamate represses the synthesis of glutamate dehydrogenase and l-glutamine represses the synthesis of glutamine synthetase. 5. A comparison of the glutamine-synthesizing activity and the γ-glutamyltransferase activity of glutamine synthetase in Aspergillus and Neurospora gave no indication that these fungi produce different forms of glutamine synthetase when grown on ammonia or l-glutamate as nitrogen sources. PMID:4901826

  16. Regulation of synthesis of glutamate dehydrogenase and glutamine synthetase in micro-organisms.

    PubMed

    Pateman, J A

    1969-12-01

    1. Aspergillus nidulans, Neurospora crassa and Escherichia coli were grown on media containing a range of concentrations of nitrate, or ammonia, or urea, or l-glutamate, or l-glutamine as the sole source of nitrogen and the glutamate dehydrogenate and glutamine synthetase of the cells measured. 2. Aspergillus, Neurospora and Escherichia coli cells, grown on l-glutamate or on high concentrations of ammonia or on high concentrations of urea, possessed low glutamate dehydrogenase activity compared with cells grown on other nitrogen sources. 3. Aspergillus, Neurospora and Escherichia coli cells grown on l-glutamate possessed high glutamine synthetase activity compared with cells grown on other nitrogen sources. 4. The hypothesis is proposed that in Aspergillus, Neurospora and Escherichia colil-glutamate represses the synthesis of glutamate dehydrogenase and l-glutamine represses the synthesis of glutamine synthetase. 5. A comparison of the glutamine-synthesizing activity and the gamma-glutamyltransferase activity of glutamine synthetase in Aspergillus and Neurospora gave no indication that these fungi produce different forms of glutamine synthetase when grown on ammonia or l-glutamate as nitrogen sources.

  17. The yeast VAS1 gene encodes both mitochondrial and cytoplasmic valyl-tRNA synthetases.

    PubMed

    Chatton, B; Walter, P; Ebel, J P; Lacroute, F; Fasiolo, F

    1988-01-05

    S1 mapping on the VAS1 structural gene indicates the existence of two classes of transcripts initiating at distinct in-frame translation start codons. The longer class of VAS1 transcripts initiates upstream of both ATG codons located 138 base pairs away and the shorter class downstream of the first ATG. A mutation that destroys the first AUG on the long message results in respiratory deficiency but does not affect viability. Mutation of the ATG at position 139 leads to lethality because the initiating methionine codon of the essential cytoplasmic valyl-tRNA synthetase has been destroyed. N-terminal protein sequence data further confirm translation initiation at ATG-139 for the cytoplasmic valyl-tRNA synthetase. From these results, we conclude that the VAS1 single gene encodes both mitochondrial and cytoplasmic valyl-tRNA synthetases. The presequence of the mitochondrial valyl-tRNA synthetase shows amino acid composition but not the amphiphilic character of imported mitochondrial proteins. From mutagenesis of the ATG-139 we conclude that the presequence specifically targets the cytoplasmically synthesized mitochondrial valyl-tRNA synthetase to the mitochondrial outer membrane and prevents binding of the enzyme core to cytoplasmic tRNAVal.

  18. Structure of the prolyl-tRNA synthetase from the eukaryotic pathogen Giardia lamblia

    SciTech Connect

    Larson, Eric T.; Kim, Jessica E.; Napuli, Alberto J.; Verlinde, Christophe L. M. J.; Fan, Erkang; Zucker, Frank H.; Van Voorhis, Wesley C.; Buckner, Frederick S.; Hol, Wim G. J.; Merritt, Ethan A.

    2012-09-01

    The structure of Giardia prolyl-tRNA synthetase cocrystallized with proline and ATP shows evidence for half-of-the-sites activity, leading to a corresponding mixture of reaction substrates and product (prolyl-AMP) in the two active sites of the dimer. The genome of the human intestinal parasite Giardia lamblia contains only a single aminoacyl-tRNA synthetase gene for each amino acid. The Giardia prolyl-tRNA synthetase gene product was originally misidentified as a dual-specificity Pro/Cys enzyme, in part owing to its unexpectedly high off-target activation of cysteine, but is now believed to be a normal representative of the class of archaeal/eukaryotic prolyl-tRNA synthetases. The 2.2 Å resolution crystal structure of the G. lamblia enzyme presented here is thus the first structure determination of a prolyl-tRNA synthetase from a eukaryote. The relative occupancies of substrate (proline) and product (prolyl-AMP) in the active site are consistent with half-of-the-sites reactivity, as is the observed biphasic thermal denaturation curve for the protein in the presence of proline and MgATP. However, no corresponding induced asymmetry is evident in the structure of the protein. No thermal stabilization is observed in the presence of cysteine and ATP. The implied low affinity for the off-target activation product cysteinyl-AMP suggests that translational fidelity in Giardia is aided by the rapid release of misactivated cysteine.

  19. Interdomain and Intermodule Organization in Epimerization Domain Containing Nonribosomal Peptide Synthetases.

    PubMed

    Chen, Wei-Hung; Li, Kunhua; Guntaka, Naga Sandhya; Bruner, Steven D

    2016-08-19

    Nonribosomal peptide synthetases are large, complex multidomain enzymes responsible for the biosynthesis of a wide range of peptidic natural products. Inherent to synthetase chemistry is the thioester templated mechanism that relies on protein/protein interactions and interdomain dynamics. Several questions related to structure and mechanism remain to be addressed, including the incorporation of accessory domains and intermodule interactions. The inclusion of nonproteinogenic d-amino acids into peptide frameworks is a common and important modification for bioactive nonribosomal peptides. Epimerization domains, embedded in nonribosomal peptide synthetases assembly lines, catalyze the l- to d-amino acid conversion. Here we report the structure of the epimerization domain/peptidyl carrier protein didomain construct from the first module of the cyclic peptide antibiotic gramicidin synthetase. Both holo (phosphopantethiene post-translationally modified) and apo structures were determined, each representing catalytically relevant conformations of the two domains. The structures provide insight into domain-domain recognition, substrate delivery during the assembly line process, in addition to the structural organization of homologous condensation domains, canonical players in all synthetase modules.

  20. Glutamine Synthetase Sensitivity to Oxidative Modification during Nutrient Starvation in Prochlorococcus marinus PCC 9511

    PubMed Central

    Gómez-Baena, Guadalupe; Domínguez-Martín, María Agustina; Donaldson, Robert P.; García-Fernández, José Manuel; Diez, Jesús

    2015-01-01

    Glutamine synthetase plays a key role in nitrogen metabolism, thus the fine regulation of this enzyme in Prochlorococcus, which is especially important in the oligotrophic oceans where this marine cyanobacterium thrives. In this work, we studied the metal-catalyzed oxidation of glutamine synthetase in cultures of Prochlorococcus marinus strain PCC 9511 subjected to nutrient limitation. Nitrogen deprivation caused glutamine synthetase to be more sensitive to metal-catalyzed oxidation (a 36% increase compared to control, non starved samples). Nutrient starvation induced also a clear increase (three-fold in the case of nitrogen) in the concentration of carbonyl derivatives in cell extracts, which was also higher (22%) upon addition of the inhibitor of electron transport, DCMU, to cultures. Our results indicate that nutrient limitations, representative of the natural conditions in the Prochlorococcus habitat, affect the response of glutamine synthetase to oxidative inactivating systems. Implications of these results on the regulation of glutamine synthetase by oxidative alteration prior to degradation of the enzyme in Prochlorococcus are discussed. PMID:26270653

  1. Cysteinyl-tRNA synthetase is not essential for viability of the archaeon Methanococcus maripaludis

    PubMed Central

    Stathopoulos, Constantinos; Kim, Wonduck; Li, Tong; Anderson, Iain; Deutsch, Britta; Palioura, Sotiria; Whitman, William; Söll, Dieter

    2001-01-01

    The methanogenic archaea Methanocaldococcus jannaschii and Methanothermobacter thermautotrophicus contain a dual-specificity prolyl-tRNA synthetase (ProCysRS) that accurately forms both prolyl-tRNA (Pro-tRNA) and cysteinyl-tRNA (Cys-tRNA) suitable for in vivo translation. This intriguing enzyme may even perform its dual role in organisms that possess a canonical single-specificity cysteinyl-tRNA synthetase (CysRS), raising the question as to whether this latter aminoacyl-tRNA synthetase is indeed required for cell viability. To test the postulate that all synthetase genes are essential, we disrupted the cysS gene (encoding CysRS) of Methanococcus maripaludis. The knockout strain was viable under normal growth conditions. Biochemical analysis showed that the pure M. maripaludis ProCysRS was capable of forming Cys-tRNA, implying that the dual-specificity enzyme compensates in vivo for the loss of CysRS. The canonical CysRS has a higher affinity for cysteine than ProCysRS, a reason why M. maripaludis may have acquired cysS by a late lateral gene transfer. These data challenge the notion that all twenty aminoacyl-tRNA synthetases are essential for the viability of a cell. PMID:11717392

  2. Glutamine Synthetase Sensitivity to Oxidative Modification during Nutrient Starvation in Prochlorococcus marinus PCC 9511.

    PubMed

    Gómez-Baena, Guadalupe; Domínguez-Martín, María Agustina; Donaldson, Robert P; García-Fernández, José Manuel; Diez, Jesús

    2015-01-01

    Glutamine synthetase plays a key role in nitrogen metabolism, thus the fine regulation of this enzyme in Prochlorococcus, which is especially important in the oligotrophic oceans where this marine cyanobacterium thrives. In this work, we studied the metal-catalyzed oxidation of glutamine synthetase in cultures of Prochlorococcus marinus strain PCC 9511 subjected to nutrient limitation. Nitrogen deprivation caused glutamine synthetase to be more sensitive to metal-catalyzed oxidation (a 36% increase compared to control, non starved samples). Nutrient starvation induced also a clear increase (three-fold in the case of nitrogen) in the concentration of carbonyl derivatives in cell extracts, which was also higher (22%) upon addition of the inhibitor of electron transport, DCMU, to cultures. Our results indicate that nutrient limitations, representative of the natural conditions in the Prochlorococcus habitat, affect the response of glutamine synthetase to oxidative inactivating systems. Implications of these results on the regulation of glutamine synthetase by oxidative alteration prior to degradation of the enzyme in Prochlorococcus are discussed.

  3. Glutamate-Dependent Translational Control of Glutamine Synthetase in Bergmann Glia Cells.

    PubMed

    Tiburcio-Félix, Reynaldo; Escalante-López, Miguel; López-Bayghen, Bruno; Martínez, Daniel; Hernández-Kelly, Luisa C; Zinker, Samuel; Hernández-Melchor, Dinorah; López-Bayghen, Esther; Olivares-Bañuelos, Tatiana N; Ortega, Arturo

    2017-09-05

    Glutamate is the major excitatory transmitter of the vertebrate brain. It exerts its actions through the activation of specific plasma membrane receptors expressed both in neurons and in glial cells. Recent evidence has shown that glutamate uptake systems, particularly enriched in glia cells, trigger biochemical cascades in a similar fashion as receptors. A tight regulation of glutamate extracellular levels prevents neuronal overstimulation and cell death, and it is critically involved in glutamate turnover. Glial glutamate transporters are responsible of the majority of the brain glutamate uptake activity. Once internalized, this excitatory amino acid is rapidly metabolized to glutamine via the astrocyte-enriched enzyme glutamine synthetase. A coupling between glutamate uptake and glutamine synthesis and release has been commonly known as the glutamate/glutamine shuttle. Taking advantage of the established model of cultured Bergmann glia cells, in this contribution, we explored the gene expression regulation of glutamine synthetase. A time- and dose-dependent regulation of glutamine synthetase protein and activity levels was found. Moreover, glutamate exposure resulted in the transient shift of glutamine synthetase mRNA from the monosomal to the polysomal fraction. These results demonstrate a novel mode of glutamate-dependent glutamine synthetase regulation and strengthen the notion of an exquisite glia neuronal interaction in glutamatergic synapses.

  4. When contemporary aminoacyl-tRNA synthetases invent their cognate amino acid metabolism

    PubMed Central

    Roy, Hervé; Becker, Hubert Dominique; Reinbolt, Joseph; Kern, Daniel

    2003-01-01

    Faithful protein synthesis relies on a family of essential enzymes called aminoacyl-tRNA synthetases, assembled in a piecewise fashion. Analysis of the completed archaeal genomes reveals that all archaea that possess asparaginyl-tRNA synthetase (AsnRS) also display a second ORF encoding an AsnRS truncated from its anticodon binding-domain (AsnRS2). We show herein that Pyrococcus abyssi AsnRS2, in contrast to AsnRS, does not sustain asparaginyl-tRNAAsn synthesis but is instead capable of converting aspartic acid into asparagine. Functional analysis and complementation of an Escherichia coli asparagine auxotrophic strain show that AsnRS2 constitutes the archaeal homologue of the bacterial ammonia-dependent asparagine synthetase A (AS-A), therefore named archaeal asparagine synthetase A (AS-AR). Primary sequence- and 3D-based phylogeny shows that an archaeal AspRS ancestor originated AS-AR, which was subsequently transferred into bacteria by lateral gene transfer in which it underwent structural changes producing AS-A. This study provides evidence that a contemporary aminoacyl-tRNA synthetase can be recruited to sustain amino acid metabolism. PMID:12874385

  5. Functional Analysis of Leishmania Cyclopropane Fatty Acid Synthetase

    PubMed Central

    Oyola, Samuel O.; Evans, Krystal J.; Smith, Terry K.; Smith, Barbara A.; Hilley, James D.; Mottram, Jeremy C.; Kaye, Paul M.; Smith, Deborah F.

    2012-01-01

    The single gene encoding cyclopropane fatty acid synthetase (CFAS) is present in Leishmania infantum, L. mexicana and L. braziliensis but absent from L. major, a causative agent of cutaneous leishmaniasis. In L. infantum, usually causative agent of visceral leishmaniasis, the CFAS gene is transcribed in both insect (extracellular) and host (intracellular) stages of the parasite life cycle. Tagged CFAS protein is stably detected in intracellular L. infantum but only during the early log phase of extracellular growth, when it shows partial localisation to the endoplasmic reticulum. Lipid analyses of L. infantum wild type, CFAS null and complemented parasites detect a low abundance CFAS-dependent C19Δ fatty acid, characteristic of a cyclopropanated species, in wild type and add-back cells. Sub-cellular fractionation studies locate the C19Δ fatty acid to both ER and plasma membrane-enriched fractions. This fatty acid is not detectable in wild type L. major, although expression of the L. infantum CFAS gene in L. major generates cyclopropanated fatty acids, indicating that the substrate for this modification is present in L. major, despite the absence of the modifying enzyme. Loss of the L. infantum CFAS gene does not affect extracellular parasite growth, phagocytosis or early survival in macrophages. However, while endocytosis is also unaffected in the extracellular CFAS nulls, membrane transporter activity is defective and the null parasites are more resistant to oxidative stress. Following infection in vivo, L. infantum CFAS nulls exhibit lower parasite burdens in both the liver and spleen of susceptible hosts but it has not been possible to complement this phenotype, suggesting that loss of C19Δ fatty acid may lead to irreversible changes in cell physiology that cannot be rescued by re-expression. Aberrant cyclopropanation in L. major decreases parasite virulence but does not influence parasite tissue tropism. PMID:23251490

  6. Antimalarial Benzoxaboroles Target Plasmodium falciparum Leucyl-tRNA Synthetase

    PubMed Central

    Sonoiki, Ebere; Palencia, Andres; Guo, Denghui; Ahyong, Vida; Dong, Chen; Li, Xianfeng; Hernandez, Vincent S.; Zhang, Yong-Kang; Choi, Wai; Gut, Jiri; Legac, Jennifer; Cooper, Roland; Alley, M. R. K.; Freund, Yvonne R.; DeRisi, Joseph; Cusack, Stephen

    2016-01-01

    There is a need for new antimalarials, ideally with novel mechanisms of action. Benzoxaboroles have been shown to be active against bacteria, fungi, and trypanosomes. Therefore, we investigated the antimalarial activity and mechanism of action of 3-aminomethyl benzoxaboroles against Plasmodium falciparum. Two 3-aminomethyl compounds, AN6426 and AN8432, demonstrated good potency against cultured multidrug-resistant (W2 strain) P. falciparum (50% inhibitory concentration [IC50] of 310 nM and 490 nM, respectively) and efficacy against murine Plasmodium berghei infection when administered orally once daily for 4 days (90% effective dose [ED90], 7.4 and 16.2 mg/kg of body weight, respectively). To characterize mechanisms of action, we selected parasites with decreased drug sensitivity by culturing with stepwise increases in concentration of AN6426. Resistant clones were characterized by whole-genome sequencing. Three generations of resistant parasites had polymorphisms in the predicted editing domain of the gene encoding a P. falciparum leucyl-tRNA synthetase (LeuRS; PF3D7_0622800) and in another gene (PF3D7_1218100), which encodes a protein of unknown function. Solution of the structure of the P. falciparum LeuRS editing domain suggested key roles for mutated residues in LeuRS editing. Short incubations with AN6426 and AN8432, unlike artemisinin, caused dose-dependent inhibition of [14C]leucine incorporation by cultured wild-type, but not resistant, parasites. The growth of resistant, but not wild-type, parasites was impaired in the presence of the unnatural amino acid norvaline, consistent with a loss of LeuRS editing activity in resistant parasites. In summary, the benzoxaboroles AN6426 and AN8432 offer effective antimalarial activity and act, at least in part, against a novel target, the editing domain of P. falciparum LeuRS. PMID:27270277

  7. Functional interactions between a glutamine synthetase promoter and MYB proteins.

    PubMed

    Gómez-Maldonado, Josefa; Avila, Concepción; Torre, Fernando; Cañas, Rafael; Cánovas, Francisco M; Campbell, Malcolm M

    2004-08-01

    In Scots pine (Pinus sylvestris), ammonium assimilation is catalysed by glutamine synthetase (GS) [EC 6.3.1.2], which is encoded by two genes, PsGS1a and PsGS1b. PsGS1b is expressed in the vascular tissue throughout the plant body, where it is believed to play a role in recycling ammonium released by various facets of metabolism. The mechanisms that may underpin the transcriptional regulation of PsGS1b were explored. The PsGS1b promoter contains a region that is enriched in previously characterized cis-acting elements, known as AC elements. Pine nuclear proteins bound these AC element-rich regions in a tissue-specific manner. As previous experiments had shown that R2R3-MYB transcription factors could interact with AC elements, the capacity of the AC elements in the PsGS1b promoter to interact with MYB proteins was examined. Two MYB proteins from loblolly pine (Pinus taeda), PtMYB1 and PtMYB4, bound to the PsGS1b promoter were able to activate transcription from this promoter in yeast, arabidopsis and pine cells. Immunolocalization experiments revealed that the two MYB proteins were most abundant in cells previously shown to accumulate PsGS1b transcripts. Immunoprecipitation analysis and supershift electrophoretic mobility shift assays implicated these same two proteins in the formation of complexes between pine nuclear extracts and the PsGS1b promoter. Given that these MYB proteins were previously shown to have the capacity to activate gene expression related to lignin biosynthesis, we hypothesize that they may function to co-regulate lignification, a process that places significant demands on nitrogen recycling, and GS, the major enzyme involved in the nitrogen recycling pathway.

  8. Nonribosomal peptide synthetase biosynthetic clusters of ESKAPE pathogens.

    PubMed

    Gulick, Andrew M

    2017-08-02

    Covering: up to 2017.Natural products are important secondary metabolites produced by bacterial and fungal species that play important roles in cellular growth and signaling, nutrient acquisition, intra- and interspecies communication, and virulence. A subset of natural products is produced by nonribosomal peptide synthetases (NRPSs), a family of large, modular enzymes that function in an assembly line fashion. Because of the pharmaceutical activity of many NRPS products, much effort has gone into the exploration of their biosynthetic pathways and the diverse products they make. Many interesting NRPS pathways have been identified and characterized from both terrestrial and marine bacterial sources. Recently, several NRPS pathways in human commensal bacterial species have been identified that produce molecules with antibiotic activity, suggesting another source of interesting NRPS pathways may be the commensal and pathogenic bacteria that live on the human body. The ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) have been identified as a significant cause of human bacterial infections that are frequently multidrug resistant. The emerging resistance profile of these organisms has prompted calls from multiple international agencies to identify novel antibacterial targets and develop new approaches to treat infections from ESKAPE pathogens. Each of these species contains several NRPS biosynthetic gene clusters. While some have been well characterized and produce known natural products with important biological roles in microbial physiology, others have yet to be investigated. This review catalogs the NRPS pathways of ESKAPE pathogens. The exploration of novel NRPS products may lead to a better understanding of the chemical communication used by human pathogens and potentially to the discovery of novel therapeutic approaches.

  9. [2'-5' olygoadenylate synthetase activity in peripheral facial paralysis].

    PubMed

    Nakazato, H; Ikeda, M

    1995-03-01

    Interferons are produced in response to viral infection and play an important part in defense by their antiviral effects. An interferon-induced enzyme, 2'-5' oligoadenylate synthetase (2-5AS) also takes an important part of the system of defense against viral infections, and its activity elevates in nonspecific viral infections. This study was designed to evaluate the usefulness of examining serum 2-5AS activity and peripheral blood WBC 2-5AS (WBC 2-5AS) as diagnostic aids of viral infections that cause facial paralysis. Samples were obtained from 83 patients with Bell's palsy, 20 with Ramsay Hunt syndrome, 74 healthy individuals, and a total of 177 subjects. In 177, we measured serum 2-5AS level in 123 subjects, WBC 2-5AS level in 57, and both in 25. Serum 2-5AS levels in Bell's palsy (60 cases) ranged from 20 to 146 pmol/dl (average: 38.5). The range in Ramsay Hunt syndrome (13) was 20-333 (average: 59.0), and in healthy controls (50), it was 20-128 (average: 41.4). WBC 2-5AS level ranged from 20 to 5900 pmol/dl (average: 733.2) in Bell's palsy (23 cases), from 20-4540 (average: 1371.4) in Ramsay Hunt syndrome (7), and from 20-903 (average: 294.5) in healthy individuals (24). There were no statistically significant differences in serum 2-5AS activities. Otherwise, there was significant difference (p < 0.01) between healthy individuals and Patients with Ramsay Hunt syndrome in WBC 2-5AS activity. In Bell's palsy, 3 cases (13.0%) with markedly high WBC 2-5AS levels existed.(ABSTRACT TRUNCATED AT 250 WORDS)

  10. How does glutamine synthetase activity determine plant tolerance to ammonium?

    PubMed

    Cruz, C; Bio, A F M; Domínguez-Valdivia, M D; Aparicio-Tejo, P M; Lamsfus, C; Martins-Loução, M A

    2006-04-01

    The wide range of plant responses to ammonium nutrition can be used to study the way ammonium interferes with plant metabolism and to assess some characteristics related with ammonium tolerance by plants. In this work we investigated the hypothesis of plant tolerance to ammonium being related with the plants' capacity to maintain high levels of inorganic nitrogen assimilation in the roots. Plants of several species (Spinacia oleracea L., Lycopersicon esculentum L., Lactuca sativa L., Pisum sativum L. and Lupinus albus L.) were grown in the presence of distinct concentrations (0.5, 1.5, 3 and 6 mM) of nitrate and ammonium. The relative contributions of the activity of the key enzymes glutamine synthetase (GS; under light and dark conditions) and glutamate dehydrogenase (GDH) were determined. The main plant organs of nitrogen assimilation (root or shoot) to plant tolerance to ammonium were assessed. The results show that only plants that are able to maintain high levels of GS activity in the dark (either in leaves or in roots) and high root GDH activities accumulate equal amounts of biomass independently of the nitrogen source available to the root medium and thus are ammonium tolerant. Plant species with high GS activities in the dark coincide with those displaying a high capacity for nitrogen metabolism in the roots. Therefore, the main location of nitrogen metabolism (shoots or roots) and the levels of GS activity in the dark are an important strategy for plant ammonium tolerance. The relative contribution of each of these parameters to species tolerance to ammonium is assessed. The efficient sequestration of ammonium in roots, presumably in the vacuoles, is considered as an additional mechanism contributing to plant tolerance to ammonium nutrition.

  11. Blockade of Glutamine Synthetase Enhances Inflammatory Response in Microglial Cells

    PubMed Central

    Palmieri, Erika M.; Menga, Alessio; Lebrun, Aurore; Hooper, Douglas C.; Butterfield, D. Allan

    2017-01-01

    Abstract Aims: Microglial cells are brain-resident macrophages engaged in surveillance and maintained in a constant state of relative inactivity. However, their involvement in autoimmune diseases indicates that in pathological conditions microglia gain an inflammatory phenotype. The mechanisms underlying this change in the microglial phenotype are still unclear. Since metabolism is an important modulator of immune cell function, we focused our attention on glutamine synthetase (GS), a modulator of the response to lipopolysaccharide (LPS) activation in other cell types, which is expressed by microglia. Results: GS inhibition enhances release of inflammatory mediators of LPS-activated microglia in vitro, leading to perturbation of the redox balance and decreased viability of cocultured neurons. GS inhibition also decreases insulin-mediated glucose uptake in microglia. In vivo, microglia-specific GS ablation enhances expression of inflammatory markers upon LPS treatment. In the spinal cords from experimental autoimmune encephalomyelitis (EAE), GS expression levels and glutamine/glutamate ratios are reduced. Innovation: Recently, metabolism has been highlighted as mediator of immune cell function through the discovery of mechanisms that (behind these metabolic changes) modulate the inflammatory response. The present study shows for the first time a metabolic mechanism mediating microglial response to a proinflammatory stimulus, pointing to GS activity as a master modulator of immune cell function and thus unraveling a potential therapeutic target. Conclusions: Our study highlights a new role of GS in modulating immune response in microglia, providing insights into the pathogenic mechanisms associated with inflammation and new strategies of therapeutic intervention. Antioxid. Redox Signal. 26, 351–363. PMID:27758118

  12. Effects of polyamine biosynthesis inhibitors on S-adenosylmethionine synthetase and S-adenosylmethionine decarboxylase activities in carrot cell cultures

    Treesearch

    S.C. Minocha; R. Minocha; A. Komamine

    1991-01-01

    Changes in the activites of S-adcnosylmethionine (SAM) synthetase (methionine adenosyltransferase, EC 2.5.1.6.) and SAM decarboxylase (EC 4.1.1.50) were studied in carrot (Daucus carota) cell cultures in response to 2,4-dichlorophenoxyacetic acid (2,4-D) and several inhibitors of polyamine biosynthesis. Activity of SAM synthetase increased...

  13. Transcription factor TnrA inhibits the biosynthetic activity of glutamine synthetase in Bacillus subtilis.

    PubMed

    Fedorova, Ksenia; Kayumov, Airat; Woyda, Kathrin; Ilinskaja, Olga; Forchhammer, Karl

    2013-05-02

    The Bacillus subtilis glutamine synthetase (GS) plays a dual role in cell metabolism by functioning as catalyst and regulator. GS catalyses the ATP-dependent synthesis of glutamine from glutamate and ammonium. Under nitrogen-rich conditions, GS becomes feedback-inhibited by high intracellular glutamine levels and then binds transcription factors GlnR and TnrA, which control the genes of nitrogen assimilation. While GS-bound TnrA is no longer able to interact with DNA, GlnR-DNA binding is shown to be stimulated by GS complex formation. In this paper we show a new physiological feature of the interaction between glutamine synthetase and TnrA. The transcription factor TnrA inhibits the biosynthetic activity of glutamine synthetase in vivo and in vitro, while the GlnR protein does not affect the activity of the enzyme.

  14. Cloning and characterization of the C. elegans histidyl-tRNA synthetase gene.

    PubMed Central

    Amaar, Y G; Baillie, D L

    1993-01-01

    In this paper, we report the cloning and sequencing of the C. elegans histidyl-tRNA synthetase gene. The complete genomic sequence, and most of the cDNA sequence, of this gene is now determined. The gene size including flanking and coding regions is 2230 nucleotides long. Three small introns (45-50 bp long) are found to interrupt the open reading frame. The open reading frame translates to 523 amino acids. This putative protein sequence shows extensive homology with the human and yeast histidyl-tRNA the histidyl-tRNA synthetase gene is a single copy gene. Hence, it is very likely that it encodes both the cytoplasmic and the mitochondrial histidyl-tRNA synthetases. It is likely to be trans-spliced since it contains a trans-splice site in its 5' untranslated region. PMID:8414990

  15. Structure of a tryptophanyl-tRNA synthetase containing an iron–sulfur cluster

    PubMed Central

    Han, Gye Won; Yang, Xiang-Lei; McMullan, Daniel; Chong, Yeeting E.; Krishna, S. Sri; Rife, Christopher L.; Weekes, Dana; Brittain, Scott M.; Abdubek, Polat; Ambing, Eileen; Astakhova, Tamara; Axelrod, Herbert L.; Carlton, Dennis; Caruthers, Jonathan; Chiu, Hsiu-Ju; Clayton, Thomas; Duan, Lian; Feuerhelm, Julie; Grant, Joanna C.; Grzechnik, Slawomir K.; Jaroszewski, Lukasz; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Kumar, Abhinav; Marciano, David; Miller, Mitchell D.; Morse, Andrew T.; Nigoghossian, Edward; Okach, Linda; Paulsen, Jessica; Reyes, Ron; van den Bedem, Henry; White, Aprilfawn; Wolf, Guenter; Xu, Qingping; Hodgson, Keith O.; Wooley, John; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Elsliger, Marc-André; Schimmel, Paul; Wilson, Ian A.

    2010-01-01

    A novel aminoacyl-tRNA synthetase that contains an iron–sulfur cluster in the tRNA anticodon-binding region and efficiently charges tRNA with tryptophan has been found in Thermotoga maritima. The crystal structure of TmTrpRS (tryptophanyl-tRNA synthetase; TrpRS; EC 6.1.1.2) reveals an iron–sulfur [4Fe–­4S] cluster bound to the tRNA anticodon-binding (TAB) domain and an l-­tryptophan ligand in the active site. None of the other T. maritima aminoacyl-tRNA synthetases (AARSs) contain this [4Fe–4S] cluster-binding motif (C-x 22-C-x 6-C-x 2-C). It is speculated that the iron–sulfur cluster contributes to the stability of TmTrpRS and could play a role in the recognition of the anticodon. PMID:20944229

  16. SOLUBLE HEPATIC δ-AMINOLEVULINIC ACID SYNTHETASE: END-PRODUCT INHIBITION OF THE PARTIALLY PURIFIED ENZYME*

    PubMed Central

    Scholnick, Perry L.; Hammaker, Lydia E.; Marver, Harvey S.

    1969-01-01

    The present study confirms the existence of hepatic δ-aminolevulinic acid synthetase in the cytosol of the liver, suggests that this enzyme may be in transit to the mitochondria, and defines some of the characteristics of the partially purified enzyme. The substrate and cofactor requirements are similar to those of mitochondrial δ-aminolevulinic acid synthetase. Heme strongly inhibits the partially purified enzyme. A number of proteins that bind heme block this inhibition, which explains previous failures to demonstrate heme inhibition in crude systems. End-product inhibition of δ-aminolevulinic acid synthetase in the mitochondria may play an important role in the regulation of heme biosynthesis in eukaryotic cells. PMID:5257968

  17. Pyrrolysyl-tRNA Synthetase: an ordinary enzyme but an outstanding genetic code expansion tool

    PubMed Central

    Wan, Wei; Tharp, Jeffery M.; Liu, Wenshe R.

    2014-01-01

    The genetic incorporation of the 22nd proteinogenic amino acid, pyrolysine (Pyl) at amber codon is achieved by the action of pyrrolysyl-tRNA synthetase (PylRS) together with its cognate tRNAPyl. Unlike most aminoacyl-tRNA synthetases, PylRS displays high substrate side chain promiscuity, low selectivity toward its substrate α-amine, and low selectivity toward the anticodon of tRNAPyl. These unique but ordinary features of PylRS as an aminoacyl-tRNA synthetase allow the Pyl incorporation machinery to be easily engineered for the genetic incorporation of more than 100 non-canonical amino acids (NCAAs) or α-hydroxy acids into proteins at amber codon and the reassignment of other codons such as ochre UAA, opal UGA, and four-base AGGA codons to code NCAAs. PMID:24631543

  18. In vitro reactivation of in vivo ammonium-inactivated glutamine synthetase from Synechocystis sp. PCC 6803.

    PubMed

    Mérida, A; Candau, P; Florencio, F J

    1991-12-16

    Glutamine synthetase from Synechocystis sp. strain PCC 6803 is inactivated by ammonium addition to cells growing with nitrate as the nitrogen source. The enzyme can be reactivated in vitro by different methods such as alkaline phosphatase treatment, but not phosphodiesterase, by raising the pH of the crude extract to values higher than 8, by increasing the ionic strength of the cell-free extract, or by preincubation with organic solvents, such as 2-propanol and ethanol. These results suggest that the loss of glutamine synthetase activity promoted by ammonium involves the non-covalent binding of a phosphorylated compound to the enzyme and support previous results that rule out the existence of an adenylylation/deadenylylation system functioning in the regulation of cyanobacterial glutamine synthetase.

  19. Dynamically orthogonal field equations for stochastic flows and particle dynamics

    DTIC Science & Technology

    2011-02-01

    of the considered phenomenon. I am also grateful to the members of my thesis committee, Dennis McLaughlin, Nicholas Patrikalakis, and Carl Wunsch for...fields implies orthogonality of their spatial Fourier, Gabor , and Wavelet transforms [5], [35]. Therefore, different DO modes always contain different

  20. Orthogonal Projection in Teaching Regression and Financial Mathematics

    ERIC Educational Resources Information Center

    Kachapova, Farida; Kachapov, Ilias

    2010-01-01

    Two improvements in teaching linear regression are suggested. The first is to include the population regression model at the beginning of the topic. The second is to use a geometric approach: to interpret the regression estimate as an orthogonal projection and the estimation error as the distance (which is minimized by the projection). Linear…

  1. Wave Propagation in Isotropic Media with Two Orthogonal Fracture Sets

    NASA Astrophysics Data System (ADS)

    Shao, S.; Pyrak-Nolte, L. J.

    2016-10-01

    Orthogonal intersecting fracture sets form fracture networks that affect the hydraulic and mechanical integrity of a rock mass. Interpretation of elastic waves propagated through orthogonal fracture networks is complicated by guided modes that propagate along and between fractures, by multiple internal reflections, as well as by scattering from fracture intersections. The existence of some or all of these potentially overlapping modes depends on local stress fields that can preferentially close or open either one or both sets of fractures. In this study, an acoustic wave front imaging system was used to examine the effect of bi-axial loading conditions on acoustic wave propagation in isotropic media containing two orthogonal fracture sets. From the experimental data, orthogonal intersecting fracture sets support guided waves that depend on fracture spacing and fracture-specific stiffnesses. In addition, fracture intersections have stronger effects on propagating wave fronts than merely the superposition of the effects of two independent fractures because of energy partitioning among transmitted/reflected waves, scattered waves and guided modes. Interpretation of the properties of fractures or fracture sets from seismic measurements must consider non-uniform fracture stiffnesses within and among fracture sets, as well as considering the striking effects of fracture intersections on wave propagation.

  2. Orthogonal Projection in Teaching Regression and Financial Mathematics

    ERIC Educational Resources Information Center

    Kachapova, Farida; Kachapov, Ilias

    2010-01-01

    Two improvements in teaching linear regression are suggested. The first is to include the population regression model at the beginning of the topic. The second is to use a geometric approach: to interpret the regression estimate as an orthogonal projection and the estimation error as the distance (which is minimized by the projection). Linear…

  3. Trusted materials using orthogonal testing. 2015 Annual report

    SciTech Connect

    Van Benthem, Mark

    2015-09-01

    The purpose of this project is to prove (or disprove) that a reasonable number of simple tests can be used to provide a unique data signature for materials, changes in which could serve as a harbinger of material deviation, prompting further evaluations. The routine tests are mutually orthogonal to any currently required materials specification tests.

  4. A four-component organogel based on orthogonal chemical interactions.

    PubMed

    Luisier, Nicolas; Schenk, Kurt; Severin, Kay

    2014-09-14

    A thermoresponsive organogel was obtained by orthogonal assembly of four compounds using dynamic covalent boronate ester and imine bonds, as well as dative boron-nitrogen bonds. It is shown that the gel state can be disrupted or reinforced by chemicals which undergo exchange reactions with the gel components.

  5. Orthogonality criterion for banishing hydrino states from standard quantum mechanics

    NASA Astrophysics Data System (ADS)

    de Castro, Antonio S.

    2007-10-01

    Orthogonality criterion is used to shown in a very simple and general way that anomalous bound-state solutions for the Coulomb potential (hydrino states) do not exist as bona fide solutions of the Schr\\"{o}dinger, Klein-Gordon and Dirac equations.

  6. Deep learning classifier based on NPCA and orthogonal feature selection

    NASA Astrophysics Data System (ADS)

    Jankowski, Stanisław; Szymański, Zbigniew; Dziomin, Uladzimir; Golovko, Vladimir; Barcz, Aleksy

    2016-09-01

    In this paper the idea of deep learning classifier is developed. The effectiveness of discriminative classifier, as e.g. multilayer perceptron, support vector machine can be improved by adding the data preprocessing blocks: orthogonal feature selection (Gram-Schmidt method) and nonlinear principal component analysis. We present the case study of various structures of deep learning systems (scenarios).

  7. Digital orthogonal receiver for wideband radar based on compressed sensing

    NASA Astrophysics Data System (ADS)

    Hou, Qingkai; Liu, Yang; Chen, Zengping; Su, Shaoying

    2014-10-01

    Digital orthogonal receiver is one of the key techniques in digital receiver of soft radar, and compressed sensing is attracting more and more attention in radar signal processing. In this paper, we propose a CS digital orthogonal receiver for wideband radar which utilizes compressed sampling in the acquisition of radar raw data. In order to reconstruct complex signal from sub-sampled raw data, a novel sparse dictionary is proposed to represent the real-valued radar raw signal sparsely. Using our dictionary and CS algorithm, we can reconstruct the complex-valued radar signal from sub-sampled echoes. Compared with conventional digital orthogonal radar receiver, the architecture of receiver in this paper is more simplified and the sampling frequency of ADC is reduced sharply. At the same time, the range profile can be obtained during the reconstruction, so the matched filtering can be eliminated in the receiver. Some experiments on ISAR imaging based on simulated data prove that the phase information of radar echoes is well reserved in our orthogonal receiver and the whole design is effective for wideband radar.

  8. Application of orthogonal eigenstructure control to flight control design

    NASA Astrophysics Data System (ADS)

    Rastgaar Aagaah, M. A.; Ahmadian, M.; Southward, S. C.

    2008-03-01

    Orthogonal eigenstructure control is used for designing a control law that decouples the dynamic modes of a flying vehicle. Orthogonal eigenstructure control is a feedback control method for linear time invariant multi-input multi-output systems. This method has been recently developed by authors. The advantage of this control method over eigenstructure assignment methods is that there is no need for defining the closed-loop poles or shaping the closed-loop eigenvectors. This method eliminates the error due to the difference between achievable and desirable eigenvectors, by finding vectors orthogonal to the open-loop eigenvectors within the achievable eigenvectors set and replacing the open-loop eigenvectors with them. This method is also applicable to the systems with non-collocated actuators and sensors. Application of this method for designing a flight control law for the lateral directional dynamics of an F-18 HARV is presented, and compared to the results of an eigenstructure assignment method. In this case study, the actuators and sensors are not collocated. It is shown that the application of the orthogonal eigenstructure control results in a more significant dynamic modes decoupling in comparison to the application of the eigenstructure assignment technique.

  9. Response Surface Model Building Using Orthogonal Arrays for Computer Experiments

    NASA Technical Reports Server (NTRS)

    Unal, Resit; Braun, Robert D.; Moore, Arlene A.; Lepsch, Roger A.

    1997-01-01

    This study investigates response surface methods for computer experiments and discusses some of the approaches available. Orthogonal arrays constructed for computer experiments are studied and an example application to a technology selection and optimization study for a reusable launch vehicle is presented.

  10. A tunable diode laser emitting orthogonally polarised radiation modes

    SciTech Connect

    Fadyushin, A B; Velichansky, Vladimir L; Senkov, N V; Lukin, M D; Scully, M O; Fleischauer, M

    2002-07-31

    A new scheme of the external cavity of a diode laser is described. The scheme provides emission of two closely spaced ({Delta}{lambda} < 10{sup -3} nm) orthogonally polarised modes, which can be tuned within the gain line, with the mode interval continuously changed. (lasers)

  11. Crossover ensembles of random matrices and skew-orthogonal polynomials

    SciTech Connect

    Kumar, Santosh; Pandey, Akhilesh

    2011-08-15

    Highlights: > We study crossover ensembles of Jacobi family of random matrices. > We consider correlations for orthogonal-unitary and symplectic-unitary crossovers. > We use the method of skew-orthogonal polynomials and quaternion determinants. > We prove universality of spectral correlations in crossover ensembles. > We discuss applications to quantum conductance and communication theory problems. - Abstract: In a recent paper (S. Kumar, A. Pandey, Phys. Rev. E, 79, 2009, p. 026211) we considered Jacobi family (including Laguerre and Gaussian cases) of random matrix ensembles and reported exact solutions of crossover problems involving time-reversal symmetry breaking. In the present paper we give details of the work. We start with Dyson's Brownian motion description of random matrix ensembles and obtain universal hierarchic relations among the unfolded correlation functions. For arbitrary dimensions we derive the joint probability density (jpd) of eigenvalues for all transitions leading to unitary ensembles as equilibrium ensembles. We focus on the orthogonal-unitary and symplectic-unitary crossovers and give generic expressions for jpd of eigenvalues, two-point kernels and n-level correlation functions. This involves generalization of the theory of skew-orthogonal polynomials to crossover ensembles. We also consider crossovers in the circular ensembles to show the generality of our method. In the large dimensionality limit, correlations in spectra with arbitrary initial density are shown to be universal when expressed in terms of a rescaled symmetry breaking parameter. Applications of our crossover results to communication theory and quantum conductance problems are also briefly discussed.

  12. Velocity field calculation for non-orthogonal numerical grids

    SciTech Connect

    Flach, G. P.

    2015-03-01

    Computational grids containing cell faces that do not align with an orthogonal (e.g. Cartesian, cylindrical) coordinate system are routinely encountered in porous-medium numerical simulations. Such grids are referred to in this study as non-orthogonal grids because some cell faces are not orthogonal to a coordinate system plane (e.g. xy, yz or xz plane in Cartesian coordinates). Non-orthogonal grids are routinely encountered at the Savannah River Site in porous-medium flow simulations for Performance Assessments and groundwater flow modeling. Examples include grid lines that conform to the sloping roof of a waste tank or disposal unit in a 2D Performance Assessment simulation, and grid surfaces that conform to undulating stratigraphic surfaces in a 3D groundwater flow model. Particle tracking is routinely performed after a porous-medium numerical flow simulation to better understand the dynamics of the flow field and/or as an approximate indication of the trajectory and timing of advective solute transport. Particle tracks are computed by integrating the velocity field from cell to cell starting from designated seed (starting) positions. An accurate velocity field is required to attain accurate particle tracks. However, many numerical simulation codes report only the volumetric flowrate (e.g. PORFLOW) and/or flux (flowrate divided by area) crossing cell faces. For an orthogonal grid, the normal flux at a cell face is a component of the Darcy velocity vector in the coordinate system, and the pore velocity for particle tracking is attained by dividing by water content. For a non-orthogonal grid, the flux normal to a cell face that lies outside a coordinate plane is not a true component of velocity with respect to the coordinate system. Nonetheless, normal fluxes are often taken as Darcy velocity components, either naively or with accepted approximation. To enable accurate particle tracking or otherwise present an accurate depiction of the velocity field for a non-orthogonal

  13. Archaeal RibL: a new FAD synthetase that is air sensitive.

    PubMed

    Mashhadi, Zahra; Xu, Huimin; Grochowski, Laura L; White, Robert H

    2010-10-12

    FAD synthetases catalyze the transfer of the AMP portion of ATP to FMN to produce FAD and pyrophosphate (PP(i)). Monofunctional FAD synthetases exist in eukaryotes, while bacteria have bifunctional enzymes that catalyze both the phosphorylation of riboflavin and adenylation of FMN to produce FAD. Analyses of archaeal genomes did not reveal the presence of genes encoding either group, yet the archaea contain FAD. Our recent identification of a CTP-dependent archaeal riboflavin kinase strongly indicated the presence of a monofunctional FAD synthetase. Here we report the identification and characterization of an archaeal FAD synthetase. Methanocaldococcus jannaschii gene MJ1179 encodes a protein that is classified in the nucleotidyl transferase protein family and was previously annotated as glycerol-3-phosphate cytidylyltransferase (GCT). The MJ1179 gene was cloned and its protein product heterologously expressed in Escherichia coli. The resulting enzyme catalyzes the adenylation of FMN with ATP to produce FAD and PP(i). The MJ1179-derived protein has been designated RibL to indicate that it follows the riboflavin kinase (RibK) step in the archaeal FAD biosynthetic pathway. Aerobically isolated RibL is active only under reducing conditions. RibL was found to require divalent metals for activity, the best activity being observed with Co(2+), where the activity was 4 times greater than that with Mg(2+). Alkylation of the two conserved cysteines in the C-terminus of the protein resulted in complete inactivation. RibL was also found to catalyze cytidylation of FMN with CTP, making the modified FAD, flavin cytidine dinucleotide (FCD). Unlike other FAD synthetases, RibL does not catalyze the reverse reaction to produce FMN and ATP from FAD and PP(i). Also in contrast to other FAD synthetases, PP(i) inhibits the activity of RibL.

  14. Is hydrogen peroxide involved in the benzyl viologen-mediated in-vivo inactivation of rat liver glutamine synthetase?

    PubMed Central

    Muriana, F. J.; Ruiz-Gutierrez, V.; Relimpio, A. M.

    1993-01-01

    After benzyl viologen administration to rats, a decrease in the rat liver glutamine synthetase activity was observed. An increase in the rat liver catalase activity was found concomitantly. In combination with the catalase inhibitor aminotriazole, benzyl viologen again diminished, but markedly, the rat liver glutamine synthetase activity. Moreover, partially purified glutamine synthetase from rat liver underwent rapid inactivation upon aerobic incubation with NAD(P)H and benzyl viologen. This inactivation was prevented by catalase, which suggests that the NAD(P)H/BV2+/O2-dependent system has a role in H2O2 production. Our results suggest that H2O2 is involved in the benzyl viologen-mediated in-vivo inactivation of the rat liver glutamine synthetase. In contrast, benzyl viologen alone or in combination with aminotriazole produced a significant increase of brain glutamine synthetase. PMID:8098954

  15. Chemical bonding: the orthogonal valence-bond view.

    PubMed

    Sax, Alexander F

    2015-04-21

    Chemical bonding is the stabilization of a molecular system by charge- and spin-reorganization processes in chemical reactions. These processes are said to be local, because the number of atoms involved is very small. With multi-configurational self-consistent field (MCSCF) wave functions, these processes can be calculated, but the local information is hidden by the delocalized molecular orbitals (MO) used to construct the wave functions. The transformation of such wave functions into valence bond (VB) wave functions, which are based on localized orbitals, reveals the hidden information; this transformation is called a VB reading of MCSCF wave functions. The two-electron VB wave functions describing the Lewis electron pair that connects two atoms are frequently called covalent or neutral, suggesting that these wave functions describe an electronic situation where two electrons are never located at the same atom; such electronic situations and the wave functions describing them are called ionic. When the distance between two atoms decreases, however, every covalent VB wave function composed of non-orthogonal atomic orbitals changes its character from neutral to ionic. However, this change in the character of conventional VB wave functions is hidden by its mathematical form. Orthogonal VB wave functions composed of orthonormalized orbitals never change their character. When localized fragment orbitals are used instead of atomic orbitals, one can decide which local information is revealed and which remains hidden. In this paper, we analyze four chemical reactions by transforming the MCSCF wave functions into orthogonal VB wave functions; we show how the reactions are influenced by changing the atoms involved or by changing their local symmetry. Using orthogonal instead of non-orthogonal orbitals is not just a technical issue; it also changes the interpretation, revealing the properties of wave functions that remain otherwise undetected.

  16. Chemical Bonding: The Orthogonal Valence-Bond View

    PubMed Central

    Sax, Alexander F.

    2015-01-01

    Chemical bonding is the stabilization of a molecular system by charge- and spin-reorganization processes in chemical reactions. These processes are said to be local, because the number of atoms involved is very small. With multi-configurational self-consistent field (MCSCF) wave functions, these processes can be calculated, but the local information is hidden by the delocalized molecular orbitals (MO) used to construct the wave functions. The transformation of such wave functions into valence bond (VB) wave functions, which are based on localized orbitals, reveals the hidden information; this transformation is called a VB reading of MCSCF wave functions. The two-electron VB wave functions describing the Lewis electron pair that connects two atoms are frequently called covalent or neutral, suggesting that these wave functions describe an electronic situation where two electrons are never located at the same atom; such electronic situations and the wave functions describing them are called ionic. When the distance between two atoms decreases, however, every covalent VB wave function composed of non-orthogonal atomic orbitals changes its character from neutral to ionic. However, this change in the character of conventional VB wave functions is hidden by its mathematical form. Orthogonal VB wave functions composed of orthonormalized orbitals never change their character. When localized fragment orbitals are used instead of atomic orbitals, one can decide which local information is revealed and which remains hidden. In this paper, we analyze four chemical reactions by transforming the MCSCF wave functions into orthogonal VB wave functions; we show how the reactions are influenced by changing the atoms involved or by changing their local symmetry. Using orthogonal instead of non-orthogonal orbitals is not just a technical issue; it also changes the interpretation, revealing the properties of wave functions that remain otherwise undetected. PMID:25906476

  17. Reduced activity of glutamine synthetase in Rhodospirillum rubrum mutants lacking the adenylyltransferase GlnE.

    PubMed

    Jonsson, Anders; Nordlund, Stefan; Teixeira, Pedro Filipe

    2009-10-01

    In the nitrogen-fixing bacterium Rhodospirillum rubrum, the GlnE adenylyltransferase (encoded by glnE) catalyzes reversible adenylylation of glutamine synthetase, thereby regulating nitrogen assimilation. We have generated glnE mutant strains that are unable to adenylylate glutamine synthetase (GS). Surprisingly, the activity of GS was lower in the mutants than in the wild type, even when grown in nitrogen-fixing conditions. Our results support the proposal that R. rubrum can only cope with the absence of an adenylylation system in the presence of lowered GS expression or activity. In general terms, this report also provides further support for the central role of GS in bacterial metabolism.

  18. The binding of tyrosinyl-5'-AMP to tyrosyl-tRNA synthetase (E.coli).

    PubMed Central

    Grosse, F; Krauss, G; Kownatzki, R; Maass, G

    1979-01-01

    The binding between tyrosyl-tRNA synthetase (E.coli) and the alkylanalogue of the aminoacyladenylate, tyrosinyl-5'-AMP, has been investigated by fluorescence titrations and rapid mixing experiments. Tyrosyl-tRNA synthetase has two equivalent and independent binding sites for tyrosinyl-5'-AMP. The intrinsic binding constant is 4 x 10(7)M-1. The binding sites for tRNATyr and tyrosinyl-5'-AMP are independent of each other, the anticooperative mode of tRNA binding being preserved in the presence of tyrosinyl-5-AMP. PMID:377229

  19. Time course of the uridylylation and adenylylation states in the glutamine synthetase bicyclic cascade.

    PubMed Central

    Varón-Castellanos, R; Havsteen, B H; García-Moreno, M; Valero-Ruiz, E; Molina-Alarcón, M; García-Cánovas, F

    1993-01-01

    A kinetic analysis of the glutamine synthetase bicyclic cascade is presented. It includes the dependence on time from the onset of the reaction of both the uridylylation of Shapiro's regulatory protein and the adenylylation of the glutamine synthetase. The transient phase equations obtained allow an estimation of the time elapsed until the states of uridylylation and adenylylation reach their steady-states, and therefore an evaluation of the effective sensitivity of the system. The contribution of the uridylylation cycle to the adenylylation cycle has been studied, and an equation relating the state of adenylylation at any time to the state of uridylylation at the same instant has been derived. PMID:8104399

  20. A Survey of Glutamine Synthetase Activities in Tissues from Three Classes of Fish.

    DTIC Science & Technology

    1980-09-01

    USA-TR-88-1 NL jj 1-5II!11111112.° MCROCOPY RESOLUTION TEST CHART NATIONAL HIJ|AL) OF STANPARL AL 61 A - USAFA-TR-4O A SURVEY OF GLUTAMINE SYNTHETASE...glutamine synthetase activity is defined as the production of one pmole of y-glutamyl hydroxamate per min at 25°C. Protein was determined by the biuret method...content. P. is listed as at progein per g tissue ( biuret method); nm ± standard deviation.’ Number of specimens examined is listed in parenthesis. 3 body

  1. Optical design through optimization for rectangular apertures using freeform orthogonal polynomials: a case study

    NASA Astrophysics Data System (ADS)

    Nikolic, Milena; Benítez, Pablo; Narasimhan, Bharathwaj; Grabovickic, Dejan; Liu, Jayao; Miñano, Juan C.

    2016-07-01

    Several applications of freeform optics call for deeper analysis of systems with rectangular apertures. We study the behavior of a freeform mirror system by comparing four orthogonal polynomial surface representations through local optimization. We compare polynomials with different orthogonal areas (rectangular-circular) and different metrics (sag-gradient). Polynomials orthogonal inside a rectangle converge faster or to a better local minimum than those orthogonal inside a circle in the example considered. This is the most likely due to the loss of the good properties of orthogonality when the orthogonality area does not coincide with the surface area used.

  2. Correlation of exon 3 β-catenin mutations with glutamine synthetase staining patterns in hepatocellular adenoma and hepatocellular carcinoma.

    PubMed

    Hale, Gillian; Liu, Xinxin; Hu, Junjie; Xu, Zhong; Che, Li; Solomon, David; Tsokos, Christos; Shafizadeh, Nafis; Chen, Xin; Gill, Ryan; Kakar, Sanjay

    2016-11-01

    The current clinical practice is based on the assumption of strong correlation between diffuse glutamine synthetase expression and β-catenin activation in hepatocellular adenoma and hepatocellular carcinoma. This high correlation is based on limited data and may represent an oversimplification as glutamine synthetase staining patterns show wide variability in clinical practice. Standardized criteria for interpreting diverse glutamine synthetase patterns, and the association between each pattern and β-catenin mutations is not clearly established. This study examines the correlation between glutamine synthetase staining patterns and β-catenin mutations in 15 typical hepatocellular adenomas, 5 atypical hepatocellular neoplasms and 60 hepatocellular carcinomas. Glutamine synthetase staining was classified into one of the three patterns: (a) diffuse homogeneous: moderate-to-strong cytoplasmic staining in >90% of lesional cells, without a map-like pattern, (b) diffuse heterogeneous: moderate-to-strong staining in 50-90% of lesional cells, without a map-like pattern, and (c) patchy: moderate-to-strong staining in <50% of lesional cells (often perivascular), or weak staining irrespective of the extent, and all other staining patterns (including negative cases). Sanger sequencing of CTNNB1 exon 3 was performed in all cases. Of hepatocellular tumors with diffuse glutamine synthetase staining (homogeneous or heterogeneous), an exon 3 β-catenin mutation was detected in 33% (2/6) of typical hepatocellular adenoma, 75% (3/4) of atypical hepatocellular neoplasm and 17% (8/47) of hepatocellular carcinomas. An exon 3 mutation was also observed in 15% (2/13) of hepatocellular carcinomas with patchy glutamine synthetase staining. The results show a modest correlation between diffuse glutamine synthetase immunostaining and exon 3 β-catenin mutations in hepatocellular adenoma and hepatocellular carcinoma with discrepancy rates >50% in both hepatocellular adenoma and hepatocellular

  3. Correlation of Exon 3 β-catenin Mutations with Glutamine Synthetase Staining Patterns in Hepatocellular Adenoma and Hepatocellular Carcinoma

    PubMed Central

    Hale, Gillian; Liu, Xinxin; Hu, Junjie; Xu, Zhong; Che, Li; Solomon, David; Tsokos, Christos; Shafizadeh, Nafis; Chen, Xin; Gill, Ryan; Kakar, Sanjay

    2016-01-01

    The current clinical practice is based on the assumption of strong correlation between diffuse glutamine synthetase expression and β-catenin activation in hepatocellular adenoma and hepatocellular carcinoma. This high correlation is based on limited data, and may represent an oversimplification as glutamine synthetase staining patterns show wide variability in clinical practice. Standardized criteria for interpreting diverse glutamine synthetase patterns, and the association between each pattern and β-catenin mutations is not clearly established. This study examines the correlation between glutamine synthetase staining patterns and β-catenin mutations in 15 typical hepatocellular adenomas, 5 atypical hepatocellular neoplasms and 60 hepatocellular carcinomas. Glutamine synthetase staining was classified into one of three patterns: (a) diffuse homogeneous: moderate to strong cytoplasmic staining in more than 90% of lesional cells, without a map-like pattern, (b) diffuse heterogeneous: moderate to strong staining in 50–90% of lesional cells, without a map-like pattern, and (c) patchy: moderate to strong staining in <50% of lesional cells (often perivascular), or weak staining irrespective of extent, and all other staining patterns (including negative cases). Sanger sequencing of CTNNB1 exon 3 was performed in all cases. Of hepatocellular tumors with diffuse glutamine synthetase staining (homogeneous or heterogeneous), an exon 3 β-catenin mutation was detected in 33% (2/6) of typical hepatocellular adenoma, 75% (3/4) of atypical hepatocellular neoplasm and 17% (8/47) of hepatocellular carcinomas. An exon 3 mutation was also observed in 15% (2/13) of hepatocellular carcinomas with patchy glutamine synthetase staining. The results show a modest correlation between diffuse glutamine synthetase immunostaining and exon 3 β-catenin mutations in hepatocellular adenoma and hepatocellular carcinoma with discrepancy rates exceeding 50% in both hepatocellular adenoma and

  4. The cytoplasmic prolyl-tRNA synthetase of the malaria parasite is a dual-stage target of febrifugine and its analogs.

    PubMed

    Herman, Jonathan D; Pepper, Lauren R; Cortese, Joseph F; Estiu, Guillermina; Galinsky, Kevin; Zuzarte-Luis, Vanessa; Derbyshire, Emily R; Ribacke, Ulf; Lukens, Amanda K; Santos, Sofia A; Patel, Vishal; Clish, Clary B; Sullivan, William J; Zhou, Huihao; Bopp, Selina E; Schimmel, Paul; Lindquist, Susan; Clardy, Jon; Mota, Maria M; Keller, Tracy L; Whitman, Malcolm; Wiest, Olaf; Wirth, Dyann F; Mazitschek, Ralph

    2015-05-20

    The emergence of drug resistance is a major limitation of current antimalarials. The discovery of new druggable targets and pathways including those that are critical for multiple life cycle stages of the malaria parasite is a major goal for developing next-generation antimalarial drugs. Using an integrated chemogenomics approach that combined drug resistance selection, whole-genome sequencing, and an orthogonal yeast model, we demonstrate that the cytoplasmic prolyl-tRNA (transfer RNA) synthetase (PfcPRS) of the malaria parasite Plasmodium falciparum is a biochemical and functional target of febrifugine and its synthetic derivative halofuginone. Febrifugine is the active principle of a traditional Chinese herbal remedy for malaria. We show that treatment with febrifugine derivatives activated the amino acid starvation response in both P. falciparum and a transgenic yeast strain expressing PfcPRS. We further demonstrate in the Plasmodium berghei mouse model of malaria that halofuginol, a new halofuginone analog that we developed, is active against both liver and asexual blood stages of the malaria parasite. Halofuginol, unlike halofuginone and febrifugine, is well tolerated at efficacious doses and represents a promising lead for the development of dual-stage next-generation antimalarials.

  5. The multifunctional peptide synthetase performing the first step of penicillin biosynthesis in Penicillium chrysogenum is a 421,073 dalton protein similar to Bacillus brevis peptide antibiotic synthetases.

    PubMed Central

    Smith, D J; Earl, A J; Turner, G

    1990-01-01

    The nucleotide sequence of the Penicillium chrysogenum Oli13 acvA gene encoding delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase, which performs the first step in penicillin biosynthesis, has been determined. The acvA gene contains an open reading frame of 11,238 bp encoding a protein of 3746 amino acids with a predicted mol. wt of 421,073 dalton. Three domains within the protein of approximately 570 amino acids have between 38% and 43% identity with each other and share similarity with two antibiotic peptide synthetases from Bacillus brevis as well as two other enzymes capable of performing ATP-pyrophosphate exchange reactions. The acvA gene is located close to the pcbC gene encoding isopenicillin N synthetase, the enzyme for the second step of beta-lactam biosynthesis, and is transcribed in the opposite orientation to it. The intergenic region of 1107 bp from which the acvA and pcbC genes are divergently transcribed has also been sequenced. PMID:2118102

  6. A new mechanism of post-transfer editing by aminoacyl-tRNA synthetases: catalysis of hydrolytic reaction by bacterial-type prolyl-tRNA synthetase.

    PubMed

    Boyarshin, Konstantin S; Priss, Anastasia E; Rayevskiy, Alexsey V; Ilchenko, Mykola M; Dubey, Igor Ya; Kriklivyi, Ivan A; Yaremchuk, Anna D; Tukalo, Michael A

    2017-02-01

    Aminoacyl tRNA synthetases are enzymes that specifically attach amino acids to cognate tRNAs for use in the ribosomal stage of translation. For many aminoacyl tRNA synthetases, the required level of amino acid specificity is achieved either by specific hydrolysis of misactivated aminoacyl-adenylate intermediate (pre-transfer editing) or by hydrolysis of the mischarged aminoacyl-tRNA (post-transfer editing). To investigate the mechanism of post-transfer editing of alanine by prolyl-tRNA synthetase from the pathogenic bacteria Enterococcus faecalis, we used molecular modeling, molecular dynamic simulations, quantum mechanical (QM) calculations, site-directed mutagenesis of the enzyme, and tRNA modification. The results support a new tRNA-assisted mechanism of hydrolysis of misacylated Ala-tRNA(Pro). The most important functional element of this catalytic mechanism is the 2'-OH group of the terminal adenosine 76 of Ala-tRNA(Pro), which forms an intramolecular hydrogen bond with the carbonyl group of the alanine residue, strongly facilitating hydrolysis. Hydrolysis was shown by QM methods to proceed via a general acid-base catalysis mechanism involving two functionally distinct water molecules. The transition state of the reaction was identified. Amino acid residues of the editing active site participate in the coordination of substrate and both attacking and assisting water molecules, performing the proton transfer to the 3'-O atom of A76.

  7. Leucyl-tRNA synthetase: double duty in amino acid sensing.

    PubMed

    Durán, Raúl V; Hall, Michael N

    2012-08-01

    The cellular response to amino acids is controlled at the molecular level by TORC1. While many of the elements that participate in TORC1 signaling are known, we still have no clear idea how cells sense amino acids. Two recent studies found that leucyl-tRNA synthetase (LRS) is a leucine sensor for TORC1, in both yeast and mammalian cells.

  8. Assembly of Multi-tRNA Synthetase Complex via Heterotetrameric Glutathione Transferase-homology Domains*

    PubMed Central

    Cho, Ha Yeon; Maeng, Seo Jin; Cho, Hyo Je; Choi, Yoon Seo; Chung, Jeong Min; Lee, Sangmin; Kim, Hoi Kyoung; Kim, Jong Hyun; Eom, Chi-Yong; Kim, Yeon-Gil; Guo, Min; Jung, Hyun Suk; Kang, Beom Sik; Kim, Sunghoon

    2015-01-01

    Many multicomponent protein complexes mediating diverse cellular processes are assembled through scaffolds with specialized protein interaction modules. The multi-tRNA synthetase complex (MSC), consisting of nine different aminoacyl-tRNA synthetases and three non-enzymatic factors (AIMP1–3), serves as a hub for many signaling pathways in addition to its role in protein synthesis. However, the assembly process and structural arrangement of the MSC components are not well understood. Here we show the heterotetrameric complex structure of the glutathione transferase (GST) domains shared among the four MSC components, methionyl-tRNA synthetase (MRS), glutaminyl-prolyl-tRNA synthetase (EPRS), AIMP2 and AIMP3. The MRS-AIMP3 and EPRS-AIMP2 using interface 1 are bridged via interface 2 of AIMP3 and EPRS to generate a unique linear complex of MRS-AIMP3:EPRS-AIMP2 at the molar ratio of (1:1):(1:1). Interestingly, the affinity at interface 2 of AIMP3:EPRS can be varied depending on the occupancy of interface 1, suggesting the dynamic nature of the linear GST tetramer. The four components are optimally arranged for maximal accommodation of additional domains and proteins. These characteristics suggest the GST tetramer as a unique and dynamic structural platform from which the MSC components are assembled. Considering prevalence of the GST-like domains, this tetramer can also provide a tool for the communication of the MSC with other GST-containing cellular factors. PMID:26472928

  9. Sponge OAS has a distinct genomic structure within the 2-5A synthetase family.

    PubMed

    Reintamm, Tõnu; Kuusksalu, Anne; Metsis, Madis; Päri, Mailis; Vallmann, Kerli; Lopp, Annika; Justesen, Just; Kelve, Merike

    2008-11-01

    2',5'-Oligoadenylate synthetases (2-5A synthetases, OAS) are enzymes that play an important role in the interferon-induced antiviral defense mechanisms in mammals. Sponges, the evolutionarily lowest multicellular animals, also possess OAS; however, their function is presently unclear. Low homology between primary structures of 2-5A synthetases from vertebrates and sponges renders their evolutionary relationship obscure. The genomic structure of vertebrate OASs has been thoroughly examined, making it possible to elucidate molecular evolution and expansion of this gene family. Until now, no OAS gene structure was available from sponges to compare it with the corresponding genes from higher organisms. In the present work, we determined the exon/intron structure of the OAS gene from the marine sponge Geodia cydonium and found it to be completely different from the strictly conserved exon/intron pattern of the OAS genes from vertebrates. This finding was corroborated by the analysis of OAS genes from another sponge, Amphimedon queenslandica, whose genome was recently sequenced. Our data suggest that vertebrate and sponge OAS genes have no direct common intron-containing ancestor and two (sub)types of OAS may be discriminated. This study opens new perspectives for understanding the phylogenesis and evolution of 2-5A synthetases as well as functional aspects of this multigene family.

  10. A decrease in S-adenosylmethionine synthetase activity increases the probability of spontaneous sporulation.

    PubMed Central

    Ochi, K; Freese, E

    1982-01-01

    Starting with a relaxed (relA) strain, mutants with reduced activity of adenosine triphosphate:L-methionine S-adenosyl transferase (EC 2.5.1.6; SAM synthetase) were isolated in Bacillus subtilis. One such mutant (gene symbol metE1) had only 3% of the normal SAM synthetase activity but grew almost as well as the parent strain. Another mutant was isolated (gene symbol spdC1) as being able to sporulate continually at a high frequency; it had one-half the normal SAM synthetase activity at 33 degrees C. Both mutants continually and spontaneously entered spore development at a higher frequency than the parent strain in a medium containing excess glucose, ammonium ions, and phosphate. Sporulation was prevented by a high concentration of SAM (1 mM or more) or by the combination of adenosine and methionine (0.5 mM or more each), both of which are precursors of SAM. In contrast to this continual increase in the spore titer, addition of decoyinine, an inhibitor of GMP synthetase, rapidly initiated massive sporulation. Various amino acid analogs also induced sporulation in the relA strain, the methionine analogs ethionine and selenomethionine being most effective. PMID:6811558

  11. Activation of chitin synthetase in permeabilized cells of a Saccharomyces cerevisiae mutant lacking proteinase B.

    PubMed Central

    Fernandez, M P; Correa, J U; Cabib, E

    1982-01-01

    Digitonin treatment at 30 degrees C of a Saccharomyces cerevisiae mutant lacking proteinase B permeabilized the cells and caused rapid and extensive activation of chitin synthetase in situ. The same result was obtained with a mutant generally defective in vacuolar proteases. By lowering the temperature and using different permeabilization procedures, we showed that increases in permeability and activation are distinct processes. Activation was inhibited by the protease inhibitors antipain and leupeptin, but by pepstatin or chymostatin. Metal chelators were also inhibitory, and their effect was reversed by the addition of Ca2+ but not by Mg2+. Antipain added together with Ca2+ after incubation of the cells in the presence of a chelating agent prevented reversal of inhibition, a result that was interpreted as indicating that antipain acts either on the same step affected by Ca2+ or on a subsequent step. Efforts to obtain activation in cell-free extracts were unsuccessful, but it was possible to extract the synthetase, once activated, by breaking permeabilized cells with glass beads. Treatment of the cell-free extracts with trypsin led not only to increased activity of chitin synthetase, but also to a change in the pH-activity curve and a diminished requirement by the enzyme for free N-acetylglucosamine. These observations suggest that the modification undergone by the synthetase during endogenous activation is different from that brought about by trypsin treatment. Images PMID:6216245

  12. A novel therapeutic target for peripheral nerve injury-related diseases: aminoacyl-tRNA synthetases

    PubMed Central

    Park, Byung Sun; Yeo, Seung Geun; Jung, Junyang; Jeong, Na Young

    2015-01-01

    Aminoacyl-tRNA synthetases (AminoARSs) are essential enzymes that perform the first step of protein synthesis. Beyond their original roles, AminoARSs possess non-canonical functions, such as cell cycle regulation and signal transduction. Therefore, AminoARSs represent a powerful pharmaceutical target if their non-canonical functions can be controlled. Using AminoARSs-specific primers, we screened mRNA expression in the spinal cord dorsal horn of rats with peripheral nerve injury created by sciatic nerve axotomy. Of 20 AminoARSs, we found that phenylalanyl-tRNA synthetase beta chain (FARSB), isoleucyl-tRNA synthetase (IARS) and methionyl-tRNA synthetase (MARS) mRNA expression was increased in spinal dorsal horn neurons on the injured side, but not in glial cells. These findings suggest the possibility that FARSB, IARS and MARS, as a neurotransmitter, may transfer abnormal sensory signals after peripheral nerve damage and become a new target for drug treatment. PMID:26692865

  13. Isolation and characterization of acetyl-coenzyme A synthetase from Methanothrix soehngenii.

    PubMed Central

    Jetten, M S; Stams, A J; Zehnder, A J

    1989-01-01

    In Methanothrix soehngenii, acetate is activated to acetyl-coenzyme A (acetyl-CoA) by an acetyl-CoA synthetase. Cell extracts contained high activities of adenylate kinase and pyrophosphatase, but no activities of a pyrophosphate:AMP and pyrophosphate:ADP phosphotransferase, indicating that the activation of 1 acetate in Methanothrix requires 2 ATP. Acetyl-CoA synthetase was purified 22-fold in four steps to apparent homogeneity. The native molecular mass of the enzyme from M. soehngenii estimated by gel filtration was 148 kilodaltons (kDa). The enzyme was composed of two subunits with a molecular mass of 73 kDa in an alpha 2 oligomeric structure. The acetyl-CoA synthetase constituted up to 4% of the soluble cell protein. At the optimum pH of 8.5, the Vmax was 55 mumol of acetyl-CoA formed per min per mg of protein. Analysis of enzyme kinetic properties revealed a Km of 0.86 mM for acetate and 48 microM for coenzyme A. With varying amounts of ATP, weak sigmoidal kinetic was observed. The Hill plot gave a slope of 1.58 +/- 0.12, suggesting two interacting substrate sites for the ATP. The kinetic properties of the acetyl-CoA synthetase can explain the high affinity for acetate of Methanothrix soehngenii. Images PMID:2571608

  14. Ammonia Fixation via Glutamine Synthetase and Glutamate Synthase in the CAM Plant Cissus quadrangularis L. 1

    PubMed Central

    Berger, Michael G.; Sprengart, Michael L.; Kusnan, Misri; Fock, Heinrich P.

    1986-01-01

    Succulent stems of Cissus quadrangularis L. (Vitaceae) contain glutamine synthetase, glutamate synthase, and glutamate dehydrogenase. The CO2 and water gas exchanges of detached internodes were typical for Crassulacean acid metabolism plants. During three physiological phases, e.g. in the dark, in the early illumination period after stomata closure, and during the late light phase with the stomata wide open, 15NH4Cl was injected into the central pith of stem sections. The kinetics of 15N labeling in glutamate and glutamine suggested that glutamine synthetase was involved in the initial ammonia fixation. In the presence of methionine sulfoximine, an inhibitor of glutamine synthetase, the incorporation of 15N derived from 15NH4Cl was almost completely inhibited. Injections of amido-15N glutamine demonstrated a potential for 15N transfer from the amido group of glutamine into glutamate which was suppressed by the glutamate synthase inhibitor, azaserine. The evidence indicates that glutamine synthetase and glutamate synthase could assimilate ammonia and cycle nitrogen during all phases of Crassulacean acid metabolism. PMID:16664820

  15. Ammonia Fixation via Glutamine Synthetase and Glutamate Synthase in the CAM Plant Cissus quadrangularis L.

    PubMed

    Berger, M G; Sprengart, M L; Kusnan, M; Fock, H P

    1986-06-01

    Succulent stems of Cissus quadrangularis L. (Vitaceae) contain glutamine synthetase, glutamate synthase, and glutamate dehydrogenase. The CO(2) and water gas exchanges of detached internodes were typical for Crassulacean acid metabolism plants. During three physiological phases, e.g. in the dark, in the early illumination period after stomata closure, and during the late light phase with the stomata wide open, (15)NH(4)Cl was injected into the central pith of stem sections. The kinetics of (15)N labeling in glutamate and glutamine suggested that glutamine synthetase was involved in the initial ammonia fixation. In the presence of methionine sulfoximine, an inhibitor of glutamine synthetase, the incorporation of (15)N derived from (15)NH(4)Cl was almost completely inhibited. Injections of amido-(15)N glutamine demonstrated a potential for (15)N transfer from the amido group of glutamine into glutamate which was suppressed by the glutamate synthase inhibitor, azaserine. The evidence indicates that glutamine synthetase and glutamate synthase could assimilate ammonia and cycle nitrogen during all phases of Crassulacean acid metabolism.

  16. Nucleotide synthetase ribozymes may have emerged first in the RNA world

    PubMed Central

    Ma, Wentao; Yu, Chunwu; Zhang, Wentao; Hu, Jiming

    2007-01-01

    Though the “RNA world” hypothesis has gained a central role in ideas concerning the origin of life, the scenario concerning its emergence remains uncertain. It has been speculated that the first scene may have been the emergence of a template-dependent RNA synthetase ribozyme, which catalyzed its own replication: thus, “RNA replicase.” However, the speculation remains uncertain, primarily because of the large sequence length requirement of such a replicase and the lack of a convincing mechanism to ensure its self-favoring features. Instead, we propose a nucleotide synthetase ribozyme as an alternative candidate, especially considering recent experimental evidence suggesting the possibility of effective nonenzymatic template-directed synthesis of RNA. A computer simulation was conducted to support our proposal. The conditions for the emergence of the nucleotide synthetase ribozyme are discussed, based on dynamic analysis on a computer. We suggest the template-dependent RNA synthetase ribozyme emerged later, perhaps after the emergence of protocells. PMID:17878321

  17. Acetyl-CoA synthetase is a conserved regulator of autophagy and lifespan

    PubMed Central

    Mirzaei, Hamed; Longo, Valter D.

    2014-01-01

    Autophagy is essential for the maintenance of cellular homeostasis during periods of stress. Eisenberg and colleagues (Eisenberg et al., 2014) now describe the central and conserved role for acetyl-CoA synthetase in regulating lifespan in yeast and flies by a mechanism involving autophagy. PMID:24703691

  18. CDC64 Encodes Cytoplasmic Alanyl-tRNA Synthetase, Ala1p, of Saccharomyces cerevisiae

    PubMed Central

    Wrobel, Carolyn; Schmidt, Emmett V.; Polymenis, Michael

    1999-01-01

    The cdc64-1 mutation causes G1 arrest in Saccharomyces cerevisiae corresponding to a type II Start phenotype. We report that CDC64 encodes Ala1p, an alanyl-tRNA synthetase. Thus, cdc64-1 might affect charging of tRNAAla and thereby initiation of cell division. PMID:10601222

  19. Aminoacyl tRNA Synthetase Deficiency Promotes Angiogenesis via the Unfolded Protein Response Pathway

    PubMed Central

    Castranova, Daniel; Davis, Andrew E.; Lo, Brigid D.; Miller, Mayumi F.; Paukstelis, Paul J.; Swift, Matthew R.; Pham, Van N.; Torres-Vázquez, Jesús; Bell, Kameha; Shaw, Kenna M.; Kamei, Makoto; Weinstein, Brant M.

    2016-01-01

    Objective Understanding the mechanisms regulating normal and pathologic angiogenesis is of great scientific and clinical interest. In this report, we show that mutations in two different aminoacyl tRNA synthetases, threonyl tRNA synthetase (tarsy58) or isoleucyl tRNA synthetase (iarsy68), lead to similar increased branching angiogenesis in developing zebrafish. Approach and Results The Unfolded Protein Response (UPR) pathway is activated by aminoacyl tRNA synthetase deficiencies, and we show that UPR genes atf4, atf6, and xbp1, as well as the key pro-angiogenic ligand vascular endothelial growth factor (vegfaa), are all up-regulated in tarsy58 and iarsy68 mutants. Finally, we show that the PERK-ATF4 arm of the UPR pathway is necessary for both the elevated vegfaa levels and increased angiogenesis observed in tarsy58 mutants. Conclusions Our results suggest that endoplasmic reticulum (ER) stress acts as a pro-angiogenic signal via UPR pathway-dependent up-regulation of vegfaa. PMID:26821951

  20. S-Adenosylmethionine synthetase 3 is important for pollen tube growth

    USDA-ARS?s Scientific Manuscript database

    S-Adenosylmethionine is widely used in a variety of biological reactions and participates in the methionine (Met) metabolic pathway. In Arabidopsis (Arabidopsis thaliana), one of the four S-adenosylmethionine synthetase genes, METHIONINE ADENOSYLTRANSFERASE3 (MAT3), is highly expressed in pollen. He...

  1. Ligand co-crystallization of aminoacyl-tRNA synthetases from infectious disease organisms.

    PubMed

    Moen, Spencer O; Edwards, Thomas E; Dranow, David M; Clifton, Matthew C; Sankaran, Banumathi; Van Voorhis, Wesley C; Sharma, Amit; Manoil, Colin; Staker, Bart L; Myler, Peter J; Lorimer, Donald D

    2017-03-16

    Aminoacyl-tRNA synthetases (aaRSs) charge tRNAs with their cognate amino acid, an essential precursor step to loading of charged tRNAs onto the ribosome and addition of the amino acid to the growing polypeptide chain during protein synthesis. Because of this important biological function, aminoacyl-tRNA synthetases have been the focus of anti-infective drug development efforts and two aaRS inhibitors have been approved as drugs. Several researchers in the scientific community requested aminoacyl-tRNA synthetases to be targeted in the Seattle Structural Genomics Center for Infectious Disease (SSGCID) structure determination pipeline. Here we investigate thirty-one aminoacyl-tRNA synthetases from infectious disease organisms by co-crystallization in the presence of their cognate amino acid, ATP, and/or inhibitors. Crystal structures were determined for a CysRS from Borrelia burgdorferi bound to AMP, GluRS from Borrelia burgdorferi and Burkholderia thailandensis bound to glutamic acid, a TrpRS from the eukaryotic pathogen Encephalitozoon cuniculi bound to tryptophan, a HisRS from Burkholderia thailandensis bound to histidine, and a LysRS from Burkholderia thailandensis bound to lysine. Thus, the presence of ligands may promote aaRS crystallization and structure determination. Comparison with homologous structures shows conformational flexibility that appears to be a recurring theme with this enzyme class.

  2. Positive newborn screen in the biochemically normal infant of a mother with treated holocarboxylase synthetase deficiency.

    PubMed

    Nyhan, W L; Willis, M; Barshop, B A; Gangoiti, J

    2009-12-01

    Expanded programmes of newborn screening permit early diagnosis in time to prevent serious complications. These programmes have begun to detect patients who might otherwise remain asymptomatic. An additional confounding variable is the positive screen that results from maternal rather than neonatal disease. This was the case in an infant in whom elevated hydroxyisovalerylcarnitine (C(5)OH) in his newborn screen was the result of placental transfer from his mother, whose holocarboxylase synthetase deficiency was being successfully treated with biotin. The mother had been diagnosed and treated with biotin prenatally. She had no phenotypic feature of holocarboxylase synthetase deficiency, most importantly no episodes ever of acute metabolic acidosis. In the infant a repeat screen was also positive. On day 28 the infant's plasma C(5)OH carnitine was 0.05 mumol/L (normal) and urinary organic acids on day 39 were normal. The mother's excretion of 3-hydroxyisovaleric acid was 109 mmol/mol creatinine. These observations indicate that holocarboxylase synthetase deficiency is one more maternal metabolic disease which may lead to a positive screen in her unaffected newborn infant. They also make the point that holocarboxylase synthetase deficiency in an infant should be detectable in programmes of neonatal screening, which was not clear previously.

  3. Erythrocyte glutathione synthetase in 5-oxoprolinuria: kinetic studies of the mutant enzyme and detection of heterozygotes.

    PubMed

    Larsson, A; Zetterström, R; Hörnell, H; Porath, U

    1976-11-15

    The primary metabolic defect in 5-oxoprolinuria is a generalized deficiency of glutathione synthetase. The activity of this enzyme was determined in cell-free extracts of erythrocytes from patients with 5-oxoprolinuria, their parents and a sibling as well as from normal control individuals. The following activities (pkat/mg of hemoglobin) for glutathione synthetase were obtained: homozygotes mean 0.10 (range 0.07-0.12), heterozygotes mean 3.1 (range 2.8-3.7) and control individuals mean 6.1 (range 5.4-6.7). These results indicate that 5-oxoprolinuria, i.e. the defective gluthione synthetase gene(s), is transmitted by autosomal recessive inheritance. Studies of the kinetics of the low remaining activity of erythrocyte glutathione synthetase in patients with 5-oxoprolinuria failed to reveal defective affinity for glycine, gamma-glutamyl-alpha-aminobutyrate, ATP and Mg2+ ions. Furthermore, the pH optimum, time curves and temperature dependence for the mutant enzyme activity did not significantly differ from the corresponding parameters observed with normal enzyme.

  4. Brain and Liver Glutamine Synthetase of Rana catesbeiana and Rana cancrivora.

    DTIC Science & Technology

    1983-07-01

    glutamine synthetase in the liver is clear for most groups. The lungfishes (Dipnoids) do not retain urea except to avoid ammonia toxicity during...York. 11. Janssens, P.A. and Cohen, P.P. 1968. Nitrogen meta- bolism in the African lungfish . Comp. Biochem. Physiol. 24, 879-886. 9 12. Pickford, G.E

  5. Pseudomonas syringae Phytotoxins: Mode of Action, Regulation, and Biosynthesis by Peptide and Polyketide Synthetases

    PubMed Central

    Bender, Carol L.; Alarcón-Chaidez, Francisco; Gross, Dennis C.

    1999-01-01

    Coronatine, syringomycin, syringopeptin, tabtoxin, and phaseolotoxin are the most intensively studied phytotoxins of Pseudomonas syringae, and each contributes significantly to bacterial virulence in plants. Coronatine functions partly as a mimic of methyl jasmonate, a hormone synthesized by plants undergoing biological stress. Syringomycin and syringopeptin form pores in plasma membranes, a process that leads to electrolyte leakage. Tabtoxin and phaseolotoxin are strongly antimicrobial and function by inhibiting glutamine synthetase and ornithine carbamoyltransferase, respectively. Genetic analysis has revealed the mechanisms responsible for toxin biosynthesis. Coronatine biosynthesis requires the cooperation of polyketide and peptide synthetases for the assembly of the coronafacic and coronamic acid moieties, respectively. Tabtoxin is derived from the lysine biosynthetic pathway, whereas syringomycin, syringopeptin, and phaseolotoxin biosynthesis requires peptide synthetases. Activation of phytotoxin synthesis is controlled by diverse environmental factors including plant signal molecules and temperature. Genes involved in the regulation of phytotoxin synthesis have been located within the coronatine and syringomycin gene clusters; however, additional regulatory genes are required for the synthesis of these and other phytotoxins. Global regulatory genes such as gacS modulate phytotoxin production in certain pathovars, indicating the complexity of the regulatory circuits controlling phytotoxin synthesis. The coronatine and syringomycin gene clusters have been intensively characterized and show potential for constructing modified polyketides and peptides. Genetic reprogramming of peptide and polyketide synthetases has been successful, and portions of the coronatine and syringomycin gene clusters could be valuable resources in developing new antimicrobial agents. PMID:10357851

  6. Salmonella typhimurium nit Is nadE: Defective Nitrogen Utilization and Ammonia-Dependent NAD Synthetase

    PubMed Central

    Schneider, Barbara L.; Reitzer, Lawrence J.

    1998-01-01

    S. typhimurium nit mutants are defective in nitrogen assimilation, despite having normal levels of assimilatory enzymes. Complementation, enzyme assays, and genetic mapping show that nit is nadE. We present evidence that ammonia, not glutamine, is the physiological substrate for eubacterial NAD synthetases and that low activity completely accounts for the mutant phenotype. PMID:9721319

  7. Aminoacylation of tRNA in the evolution of an aminoacyl-tRNA synthetase

    PubMed Central

    Lipman, Richard S. A.; Hou, Ya-Ming

    1998-01-01

    Aminoacyl-tRNA synthetases catalyze aminoacylation of tRNAs by joining an amino acid to its cognate tRNA. The selection of the cognate tRNA is jointly determined by separate structural domains that examine different regions of the tRNA. The cysteine-tRNA synthetase of Escherichia coli has domains that select for tRNAs containing U73, the GCA anticodon, and a specific tertiary structure at the corner of the tRNA L shape. The E. coli enzyme does not efficiently recognize the yeast or human tRNACys, indicating the evolution of determinants for tRNA aminoacylation from E. coli to yeast to human and the coevolution of synthetase domains that interact with these determinants. By successively modifying the yeast and human tRNACys to ones that are efficiently aminoacylated by the E. coli enzyme, we have identified determinants of the tRNA that are important for aminoacylation but that have diverged in the course of evolution. These determinants provide clues to the divergence of synthetase domains. We propose that the domain for selecting U73 is conserved in evolution. In contrast, we propose that the domain for selecting the corner of the tRNA L shape diverged early, after the separation between E. coli and yeast, while that for selecting the GCA-containing anticodon loop diverged late, after the separation between yeast and human. PMID:9811828

  8. A NONSTEADY STATE MODEL FOR THE TIGHT-BINDING INHIBITION OF THYMIDYLATE SYNTHETASE BY 5-FLUOROURACIL

    EPA Science Inventory

    5-Fluorouracil (5_FU) is a widely used chemotherapeutic drug and tratogen that was chosen as a prototypic toxicant to contruct a biologically based dose-resonse (BBDR) model (Setzer et. al., 2001). Part of the BBDR model simulates the inhibition of thymidylate synthetase (TS), a...

  9. Tracking Lung Tumors in Orthogonal X-Rays

    PubMed Central

    2013-01-01

    This paper presents a computationally very efficient, robust, automatic tracking method that does not require any implanted fiducials for low-contrast tumors. First, it generates a set of motion hypotheses and computes corresponding feature vectors in local windows within orthogonal-axis X-ray images. Then, it fits a regression model that maps features to 3D tumor motions by minimizing geodesic distances on motion manifold. These hypotheses can be jointly generated in 3D to learn a single 3D regression model or in 2D through back projection to learn two 2D models separately. Tumor is tracked by applying regression to the consecutive image pairs while selecting optimal window size at every time. Evaluations are performed on orthogonal X-ray videos of 10 patients. Comparative experimental results demonstrate superior accuracy (~1 pixel average error) and robustness to varying imaging artifacts and noise at the same time. PMID:23986789

  10. Entanglement as a resource to distinguish orthogonal product states

    PubMed Central

    Zhang, Zhi-Chao; Gao, Fei; Cao, Tian-Qing; Qin, Su-Juan; Wen, Qiao-Yan

    2016-01-01

    It is known that there are many sets of orthogonal product states which cannot be distinguished perfectly by local operations and classical communication (LOCC). However, these discussions have left the following open question: What entanglement resources are necessary and/or sufficient for this task to be possible with LOCC? In m ⊗ n, certain classes of unextendible product bases (UPB) which can be distinguished perfectly using entanglement as a resource, had been presented in 2008. In this paper, we present protocols which use entanglement more efficiently than teleportation to distinguish some classes of orthogonal product states in m ⊗ n, which are not UPB. For the open question, our results offer rather general insight into why entanglement is useful for such tasks, and present a better understanding of the relationship between entanglement and nonlocality. PMID:27458034

  11. Entanglement as a resource to distinguish orthogonal product states.

    PubMed

    Zhang, Zhi-Chao; Gao, Fei; Cao, Tian-Qing; Qin, Su-Juan; Wen, Qiao-Yan

    2016-07-26

    It is known that there are many sets of orthogonal product states which cannot be distinguished perfectly by local operations and classical communication (LOCC). However, these discussions have left the following open question: What entanglement resources are necessary and/or sufficient for this task to be possible with LOCC? In m ⊗ n, certain classes of unextendible product bases (UPB) which can be distinguished perfectly using entanglement as a resource, had been presented in 2008. In this paper, we present protocols which use entanglement more efficiently than teleportation to distinguish some classes of orthogonal product states in m ⊗ n, which are not UPB. For the open question, our results offer rather general insight into why entanglement is useful for such tasks, and present a better understanding of the relationship between entanglement and nonlocality.

  12. Entanglement as a resource to distinguish orthogonal product states

    NASA Astrophysics Data System (ADS)

    Zhang, Zhi-Chao; Gao, Fei; Cao, Tian-Qing; Qin, Su-Juan; Wen, Qiao-Yan

    2016-07-01

    It is known that there are many sets of orthogonal product states which cannot be distinguished perfectly by local operations and classical communication (LOCC). However, these discussions have left the following open question: What entanglement resources are necessary and/or sufficient for this task to be possible with LOCC? In m ⊗ n, certain classes of unextendible product bases (UPB) which can be distinguished perfectly using entanglement as a resource, had been presented in 2008. In this paper, we present protocols which use entanglement more efficiently than teleportation to distinguish some classes of orthogonal product states in m ⊗ n, which are not UPB. For the open question, our results offer rather general insight into why entanglement is useful for such tasks, and present a better understanding of the relationship between entanglement and nonlocality.

  13. Orthogonally modulated molecular transport junctions for resettable electronic logic gates

    PubMed Central

    Meng, Fanben; Hervault, Yves-Marie; Shao, Qi; Hu, Benhui; Norel, Lucie; Rigaut, Stéphane; Chen, Xiaodong

    2014-01-01

    Individual molecules have been demonstrated to exhibit promising applications as functional components in the fabrication of computing nanocircuits. Based on their advantage in chemical tailorability, many molecular devices with advanced electronic functions have been developed, which can be further modulated by the introduction of external stimuli. Here, orthogonally modulated molecular transport junctions are achieved via chemically fabricated nanogaps functionalized with dithienylethene units bearing organometallic ruthenium fragments. The addressable and stepwise control of molecular isomerization can be repeatedly and reversibly completed with a judicious use of the orthogonal optical and electrochemical stimuli to reach the controllable switching of conductivity between two distinct states. These photo-/electro-cooperative nanodevices can be applied as resettable electronic logic gates for Boolean computing, such as a two-input OR and a three-input AND-OR. The proof-of-concept of such logic gates demonstrates the possibility to develop multifunctional molecular devices by rational chemical design. PMID:24394717

  14. Orthogonally modulated molecular transport junctions for resettable electronic logic gates

    NASA Astrophysics Data System (ADS)

    Meng, Fanben; Hervault, Yves-Marie; Shao, Qi; Hu, Benhui; Norel, Lucie; Rigaut, Stéphane; Chen, Xiaodong

    2014-01-01

    Individual molecules have been demonstrated to exhibit promising applications as functional components in the fabrication of computing nanocircuits. Based on their advantage in chemical tailorability, many molecular devices with advanced electronic functions have been developed, which can be further modulated by the introduction of external stimuli. Here, orthogonally modulated molecular transport junctions are achieved via chemically fabricated nanogaps functionalized with dithienylethene units bearing organometallic ruthenium fragments. The addressable and stepwise control of molecular isomerization can be repeatedly and reversibly completed with a judicious use of the orthogonal optical and electrochemical stimuli to reach the controllable switching of conductivity between two distinct states. These photo-/electro-cooperative nanodevices can be applied as resettable electronic logic gates for Boolean computing, such as a two-input OR and a three-input AND-OR. The proof-of-concept of such logic gates demonstrates the possibility to develop multifunctional molecular devices by rational chemical design.

  15. Design of robust differential microphone arrays with orthogonal polynomials.

    PubMed

    Pan, Chao; Benesty, Jacob; Chen, Jingdong

    2015-08-01

    Differential microphone arrays have the potential to be widely deployed in hands-free communication systems thanks to their frequency-invariant beampatterns, high directivity factors, and small apertures. Traditionally, they are designed and implemented in a multistage way with uniform linear geometries. This paper presents an approach to the design of differential microphone arrays with orthogonal polynomials, more specifically with Jacobi polynomials. It first shows how to express the beampatterns as a function of orthogonal polynomials. Then several differential beamformers are derived and their performance depends on the parameters of the Jacobi polynomials. Simulations show the great flexibility of the proposed method in terms of designing any order differential microphone arrays with different beampatterns and controlling white noise gain.

  16. Guide RNA functional modules direct Cas9 activity and orthogonality.

    PubMed

    Briner, Alexandra E; Donohoue, Paul D; Gomaa, Ahmed A; Selle, Kurt; Slorach, Euan M; Nye, Christopher H; Haurwitz, Rachel E; Beisel, Chase L; May, Andrew P; Barrangou, Rodolphe

    2014-10-23

    The RNA-guided Cas9 endonuclease specifically targets and cleaves DNA in a sequence-dependent manner and has been widely used for programmable genome editing. Cas9 activity is dependent on interactions with guide RNAs, and evolutionarily divergent Cas9 nucleases have been shown to work orthogonally. However, the molecular basis of selective Cas9:guide-RNA interactions is poorly understood. Here, we identify and characterize six conserved modules within native crRNA:tracrRNA duplexes and single guide RNAs (sgRNAs) that direct Cas9 endonuclease activity. We show the bulge and nexus are necessary for DNA cleavage and demonstrate that the nexus and hairpins are instrumental in defining orthogonality between systems. In contrast, the crRNA:tracrRNA complementary region can be modified or partially removed. Collectively, our results establish guide RNA features that drive DNA targeting by Cas9 and open new design and engineering avenues for CRISPR technologies.

  17. CVS Decomposition of 3D Homogeneous Turbulence Using Orthogonal Wavelets

    NASA Technical Reports Server (NTRS)

    Farge, Marie; Schneider, Kai; Pellegrino, Giulio; Wray, A. A.; Rogallo, R. S.

    2000-01-01

    This paper compares the filtering used in Coherent Vortex Simulation (CVS) decomposition with an orthogonal wavelet basis, with the Proper Orthogonal Decomposition (POD) or Fourier filtering. Both methods are applied to a field of Direct Numerical Simulation (DNS) data of 3D forced homogeneous isotropic turbulence at microscale Reynolds number R(sub lambda) = 168. We show that, with only 3%N retained modes, CVS filtering separates the coherent vortex tubes from the incoherent background flow. The latter is structureless, has an equipartition energy spectrum, and has a Gaussian velocity probability distribution function (PDF) and an exponential vorticity PDF. On the other hand, the Fourier basis does not extract the coherent vortex tubes cleanly and leaves organized structures in the residual high wavenumber modes whose PDFs are stretched exponentials for both the velocity and the vorticity.

  18. Force Modelling in Orthogonal Cutting Considering Flank Wear Effect

    NASA Astrophysics Data System (ADS)

    Rathod, Kanti Bhikhubhai; Lalwani, Devdas I.

    2017-05-01

    In the present work, an attempt has been made to provide a predictive cutting force model during orthogonal cutting by combining two different force models, that is, a force model for a perfectly sharp tool plus considering the effect of edge radius and a force model for a worn tool. The first force model is for a perfectly sharp tool that is based on Oxley's predictive machining theory for orthogonal cutting as the Oxley's model is for perfectly sharp tool, the effect of cutting edge radius (hone radius) is added and improve model is presented. The second force model is based on worn tool (flank wear) that was proposed by Waldorf. Further, the developed combined force model is also used to predict flank wear width using inverse approach. The performance of the developed combined total force model is compared with the previously published results for AISI 1045 and AISI 4142 materials and found reasonably good agreement.

  19. Modular inflation and the orthogonal axion as the curvaton

    SciTech Connect

    Dimopoulos, K.; Lazarides, G.

    2006-01-15

    We study a particular supersymmetric realization of the Peccei-Quinn symmetry which provides a suitable candidate for the curvaton field. The class of models considered also solves the {mu} problem, while generating the Peccei-Quinn scale dynamically. The curvaton candidate is a pseudo-Nambu-Goldstone boson corresponding to an angular degree of freedom orthogonal to the axion field. Its order parameter increases substantially following a phase transition during inflation. This results in a drastic amplification of the curvaton perturbations. Consequently, the mechanism is able to accommodate low-scale inflation with Hubble parameter at the TeV scale. Hence, we investigate modular inflation using a string axion field as the inflaton with inflation scale determined by gravity mediated soft supersymmetry breaking. We find that modular inflation with the orthogonal axion as curvaton can indeed account for the observations for natural values of the parameters.

  20. Ultrahigh-speed "orthogonal" TDM transmission with an optical Nyquist pulse train.

    PubMed

    Nakazawa, Masataka; Hirooka, Toshihiko; Ruan, Peng; Guan, Pengyu

    2012-01-16

    We propose a novel "orthogonal" TDM transmission scheme using an optical Nyquist pulse that enables us to achieve an ultrahigh data rate and spectral efficiency simultaneously without any intersymbol interference (ISI). We analytically describe the principle of orthogonal TDM, and demonstrate a 160 Gbaud optical orthogonal TDM transmission using 40 GHz optical Nyquist pulses. Tolerance to GVD and the dispersion slope is significantly improved by virtue of the orthogonality, reduced bandwidth, and minimum ISI.

  1. The effect of portacaval anastomosis on the expression of glutamine synthetase and ornithine aminotransferase in perivenous hepatocytes.

    PubMed

    da Silva, Robin; Levillain, Oliver; Brosnan, John T; Araneda, Silvia; Brosnan, Margaret E

    2013-05-01

    There is functional zonation of metabolism across the liver acinus, with glutamine synthetase restricted to a narrow band of cells around the terminal hepatic venules. Portacaval anastomosis, where there is a major rerouting of portal blood flow from the portal vein directly to the vena cava bypassing the liver, has been reported to result in a marked decrease in the activity of glutamine synthetase. It is not known whether this represents a loss of perivenous hepatocytes or whether there is a specific loss of glutamine synthetase. To answer this question, we have determined the activity of glutamine synthetase and another enzyme from the perivenous compartment, ornithine aminotransferase, as well as the immunochemical localization of both glutamine synthetase and ornithine aminotransferase in rats with a portacaval shunt. The portacaval shunt caused a marked decrease in glutamine synthetase activity and an increase in ornithine aminotransferase activity. Immunohistochemical analysis showed that the glutamine synthetase and ornithine aminotransferase proteins maintained their location in the perivenous cells. These results indicate that there is no generalized loss of perivenous hepatocytes, but rather, there is a significant alteration in the expression of these proteins and hence metabolism in this cell population.

  2. Second-order evaluations of orthogonal and symplectic Yangians

    NASA Astrophysics Data System (ADS)

    Karakhanyan, D. R.; Kirschner, R.

    2017-08-01

    Orthogonal or symplectic Yangians are defined by the Yang-Baxter RLL relation involving the fundamental R-matrix with the corresponding so( n) or sp(2 m) symmetry. We investigate the second-order solution conditions, where the expansion of L( u) in u -1 is truncated at the second power, and we derive the relations for the two nontrivial terms in L( u).

  3. A general boundary capability embedded in an orthogonal mesh

    SciTech Connect

    Hewett, D.W.; Yu-Jiuan Chen

    1995-07-01

    The authors describe how they hold onto orthogonal mesh discretization when dealing with curved boundaries. Special difference operators were constructed to approximate numerical zones split by the domain boundary; the operators are particularly simple for this rectangular mesh. The authors demonstrated that this simple numerical approach, termed Dynamic Alternating Direction Implicit, turned out to be considerably more efficient than more complex grid-adaptive algorithms that were tried previously.

  4. Orthogonal Fabry-Pérot sensors for photoacoustic tomography

    NASA Astrophysics Data System (ADS)

    Ellwood, R.; Ogunlade, O.; Zhang, E. Z.; Beard, P. C.; Cox, B. T.

    2016-03-01

    Fabry-Pérot (FP) sensors have been used to produce in-vivo photoacoustic images of exquisite quality. However, for simplicity of construction FP sensors are produced in a planar form. Planar sensors suffer from a limited detection aperture, due to their planarity. We present a novel sensor geometry that allowed a greater field of view by placing a second sensor orthogonal to the first. This captured data from the deeper lying regions of interest and mitigated the limited view.

  5. Improved Efficient, Nearly Orthogonal, Nearly Balanced Mixed Designs

    DTIC Science & Technology

    2011-12-01

    been widely used for computational experiments ( Sacks et al. 1989). They tend to have good space-filling and orthogonality behavior if n p, but when n...Fisher, R. A. 2000. Statistical methods for research workers. Edinburgh, Scotland: Oliver and Boyd. Hedayat, A. S., N. J. Sloane, and J. Stufken. 1999...methods. 2nd ed. Hoboken, New Jersey: Wiley. Sacks , J., W. J. Welch, T. J. Mitchell, and H. P. Wynn. 1989. Design and analysis of computer experiments

  6. Orthogonal mapping: A computer program for quantifying shape differences

    USGS Publications Warehouse

    Huffman, T.; Christopher, R.A.; Hazel, J.E.

    1978-01-01

    Quantitative differences in the shape of two or more objects can be obtained by Orthogonal Mapping provided coincidental or analogous points can be identified on each object. The least-squares difference between each set of analogous points is determined by use of a projective transformation of a set of measured points which involves the rotation, translocation, and scaling of these points relative to a set of fixed points. ?? 1978.

  7. Using Non-Orthogonal Iris Images for Iris Recognition

    DTIC Science & Technology

    2006-05-05

    Figure 4. Rectangular-to-polar coordinate transformation. glare and eyelashes are then accounted for by determining if any...Coordinate Transformation Boundary of pupil/iris Lower eyelid & eyelashes Center of pupil Upper eyelid & eyelashes glare Image in polar coordinates 130...are not hidden by glare, eyelids, and eyelashes . In the case of non-orthogonal iris images, the pupillary and limbic boundaries are now elliptical

  8. The Sedimentation of Particles under Orthogonal Shear in Viscoelastic Fluids

    NASA Astrophysics Data System (ADS)

    Murch, William L.; Krishnan, Sreenath; Shaqfeh, Eric S. G.

    2016-11-01

    Many engineering applications, including oil and gas recovery, require the suspension of particles in viscoelastic fluids during fluid transport and processing. A topic of specific importance involves such particle suspensions experiencing an applied shear flow in a direction perpendicular to gravity (referred to as orthogonal shear). Previously, it has been shown that particle sedimentation coupled with an orthogonal shear flow can reduce the particle settling rate in elastic fluids. The underlying mechanism of this enhanced coupling drag is not fully understood, particularly at finite Weissenberg numbers. This talk examines the role of fluid elasticity on a single, non-Brownian, rigid sphere settling in orthogonal shear using experiments and numerical simulations. New experiments were performed in a Taylor-Couette flow cell using Boger fluids to study the coupling drag as a function of the shear and sedimentation Weissenberg numbers as well as particle confinement. The elastic effect was also studied with fully 3D simulations of flow past a rigid sphere, using the FENE-P constitutive model to describe the polymeric fluid rheology. These simulations show good agreement with the experiments and allow for further insight into the mechanism of elasticity-enhanced drag. This material is based upon work supported by the National Science Foundation Graduate Research Fellowship.

  9. Parallel and orthogonal stimulus in ultradiluted neural networks.

    PubMed

    Sobral, G A; Vieira, V M; Lyra, M L; da Silva, C R

    2006-10-01

    Extending a model due to Derrida, Gardner, and Zippelius, we have studied the recognition ability of an extreme and asymmetrically diluted version of the Hopfield model for associative memory by including the effect of a stimulus in the dynamics of the system. We obtain exact results for the dynamic evolution of the average network superposition. The stimulus field was considered as proportional to the overlapping of the state of the system with a particular stimulated pattern. Two situations were analyzed, namely, the external stimulus acting on the initialization pattern (parallel stimulus) and the external stimulus acting on a pattern orthogonal to the initialization one (orthogonal stimulus). In both cases, we obtained the complete phase diagram in the parameter space composed of the stimulus field, thermal noise, and network capacity. Our results show that the system improves its recognition ability for parallel stimulus. For orthogonal stimulus two recognition phases emerge with the system locking at the initialization or stimulated pattern. We confront our analytical results with numerical simulations for the noiseless case T = 0.

  10. Animation of orthogonal texture patterns for vector field visualization.

    PubMed

    Bachthaler, Sven; Weiskopf, Daniel

    2008-01-01

    This paper introduces orthogonal vector field visualization on 2D manifolds: a representation by lines that are perpendicular to the input vector field. Line patterns are generated by line integral convolution (LIC). This visualization is combined with animation based on motion along the vector field. This decoupling of the line direction from the direction of animation allows us to choose the spatial frequencies along the direction of motion independently from the length scales along the LIC line patterns. Vision research indicates that local motion detectors are tuned to certain spatial frequencies of textures, and the above decoupling enables us to generate spatial frequencies optimized for motion perception. Furthermore, we introduce a combined visualization that employs orthogonal LIC patterns together with conventional, tangential streamline LIC patterns in order to benefit from the advantages of these two visualization approaches. In addition, a filtering process is described to achieve a consistent and temporally coherent animation of orthogonal vector field visualization. Different filter kernels and filter methods are compared and discussed in terms of visualization quality and speed. We present respective visualization algorithms for 2D planar vector fields and tangential vector fields on curved surfaces, and demonstrate that those algorithms lend themselves to efficient and interactive GPU implementations.

  11. Modular and Orthogonal Synthesis of Hybrid Polymers and Networks

    PubMed Central

    Liu, Shuang; Dicker, Kevin T.; Jia, Xinqiao

    2015-01-01

    Biomaterials scientists strive to develop polymeric materials with distinct chemical make-up, complex molecular architectures, robust mechanical properties and defined biological functions by drawing inspirations from biological systems. Salient features of biological designs include (1) repetitive presentation of basic motifs; and (2) efficient integration of diverse building blocks. Thus, an appealing approach to biomaterials synthesis is to combine synthetic and natural building blocks in a modular fashion employing novel chemical methods. Over the past decade, orthogonal chemistries have become powerful enabling tools for the modular synthesis of advanced biomaterials. These reactions require building blocks with complementary functionalities, occur under mild conditions in the presence of biological molecules and living cells and proceed with high yield and exceptional selectivity. These chemistries have facilitated the construction of complex polymers and networks in a step-growth fashion, allowing facile modulation of materials properties by simple variations of the building blocks. In this review, we first summarize features of several types of orthogonal chemistries. We then discuss recent progress in the synthesis of step growth linear polymers, dendrimers and networks that find application in drug delivery, 3D cell culture and tissue engineering. Overall, orthogonal reactions and modulular synthesis have not only minimized the steps needed for the desired chemical transformations but also maximized the diversity and functionality of the final products. The modular nature of the design, combined with the potential synergistic effect of the hybrid system, will likely result in novel hydrogel matrices with robust structures and defined functions. PMID:25572255

  12. Regression Analysis of Correlated Ordinal Data Using Orthogonalized Residuals

    PubMed Central

    Perin, J.; Preisser, J. S.; Phillips, C.; Qaqish, B.

    2015-01-01

    Summary Semi-parametric regression models for the joint estimation of marginal mean and within-cluster pairwise association parameters are used in a variety of settings for population-averaged modeling of multivariate categorical outcomes. Recently, a formulation of alternating logistic regressions based on orthogonalized, marginal residuals has been introduced for correlated binary data. Unlike the original procedure based on conditional residuals, its covariance estimator is invariant to the ordering of observations within clusters. In this article, the orthogonalized residuals method is extended to model correlated ordinal data with a global odds ratio, and shown in a simulation study to be more eficient and less biased with regards to estimating within-cluster association parameters than an existing extension to ordinal data of alternating logistic regressions based on conditional residuals. Orthogonalized residuals are used to estimate a model for three correlated ordinal outcomes measured repeatedly in a longitudinal clinical trial of an intervention to improve recovery of patients’ perception of altered sensation following jaw surgery. PMID:25134789

  13. Regression analysis of correlated ordinal data using orthogonalized residuals.

    PubMed

    Perin, J; Preisser, J S; Phillips, C; Qaqish, B

    2014-12-01

    Semi-parametric regression models for the joint estimation of marginal mean and within-cluster pairwise association parameters are used in a variety of settings for population-averaged modeling of multivariate categorical outcomes. Recently, a formulation of alternating logistic regressions based on orthogonalized, marginal residuals has been introduced for correlated binary data. Unlike the original procedure based on conditional residuals, its covariance estimator is invariant to the ordering of observations within clusters. In this article, the orthogonalized residuals method is extended to model correlated ordinal data with a global odds ratio, and shown in a simulation study to be more efficient and less biased with regards to estimating within-cluster association parameters than an existing extension to ordinal data of alternating logistic regressions based on conditional residuals. Orthogonalized residuals are used to estimate a model for three correlated ordinal outcomes measured repeatedly in a longitudinal clinical trial of an intervention to improve recovery of patients' perception of altered sensation following jaw surgery. © 2014, The International Biometric Society.

  14. Minimal parameter solution of the orthogonal matrix differential equation

    NASA Technical Reports Server (NTRS)

    Bar-Itzhack, Itzhack Y.; Markley, F. Landis

    1988-01-01

    As demonstrated in this work, all orthogonal matrices solve a first order differential equation. The straightforward solution of this equation requires n sup 2 integrations to obtain the element of the nth order matrix. There are, however, only n(n-1)/2 independent parameters which determine an orthogonal matrix. The questions of choosing them, finding their differential equation and expressing the orthogonal matrix in terms of these parameters are considered. Several possibilities which are based on attitude determination in three dimensions are examined. It is shown that not all 3-D methods have useful extensions to higher dimensions. It is also shown why the rate of change of the matrix elements, which are the elements of the angular rate vector in 3-D, are the elements of a tensor of the second rank (dyadic) in spaces other than three dimensional. It is proven that the 3-D Gibbs vector (or Cayley Parameters) are extendable to other dimensions. An algorithm is developed employing the resulting parameters, which are termed Extended Rodrigues Parameters, and numerical results are presented of the application of the algorithm to a fourth order matrix.

  15. Counting vectors for Orthogonal Fractional Factorial Design generation

    NASA Astrophysics Data System (ADS)

    Fontana, Roberto

    2011-11-01

    Generation of orthogonal fractional factorial designs (OFFDs) is an important and extensively studied subject in applied statistics. In this paper we present a methodology based on counting vectors and a methodology based on polynomial counting functions and strata,[1, 2, 3]. Both methodologies allow us to represent the OFFDs that satisfy a given set of constraints, expressed in terms of orthogonality between simple and interaction effects, as the positive integer solutions Y of a homogeneous system AY = 0 of linear equations. We show how to use this system AY = 0 • to compute, for smaller cases, a minimal set of generators of all the OFFDs (Hilbert basis); • to obtain, for larger cases, a sample of OFFDs. Finally we describe a method to find minimum size OFFDs. We set up an optimisation problem where the cost function to be minimized was the size of the OFFD and the constraints were represented by the system AY = 0. Then we searched for a solution using standard integer programming techniques. It is worth noting that the methodology does not put any restriction on the number of levels of each factor and so it can be applied to a very wide range of designs, including mixed orthogonal arrays.

  16. Minimal parameter solution of the orthogonal matrix differential equation

    NASA Technical Reports Server (NTRS)

    Bar-Itzhack, Itzhack Y.; Markley, F. Landis

    1990-01-01

    As demonstrated in this work, all orthogonal matrices solve a first order differential equation. The straightforward solution of this equation requires n sup 2 integrations to obtain the element of the nth order matrix. There are, however, only n(n-1)/2 independent parameters which determine an orthogonal matrix. The questions of choosing them, finding their differential equation and expressing the orthogonal matrix in terms of these parameters are considered. Several possibilities which are based on attitude determination in three dimensions are examined. It is shown that not all 3-D methods have useful extensions to higher dimensions. It is also shown why the rate of change of the matrix elements, which are the elements of the angular rate vector in 3-D, are the elements of a tensor of the second rank (dyadic) in spaces other than three dimensional. It is proven that the 3-D Gibbs vector (or Cayley Parameters) are extendable to other dimensions. An algorithm is developed emplying the resulting parameters, which are termed Extended Rodrigues Parameters, and numerical results are presented of the application of the algorithm to a fourth order matrix.

  17. Parallel and orthogonal stimulus in ultradiluted neural networks

    NASA Astrophysics Data System (ADS)

    Sobral, G. A., Jr.; Vieira, V. M.; Lyra, M. L.; da Silva, C. R.

    2006-10-01

    Extending a model due to Derrida, Gardner, and Zippelius, we have studied the recognition ability of an extreme and asymmetrically diluted version of the Hopfield model for associative memory by including the effect of a stimulus in the dynamics of the system. We obtain exact results for the dynamic evolution of the average network superposition. The stimulus field was considered as proportional to the overlapping of the state of the system with a particular stimulated pattern. Two situations were analyzed, namely, the external stimulus acting on the initialization pattern (parallel stimulus) and the external stimulus acting on a pattern orthogonal to the initialization one (orthogonal stimulus). In both cases, we obtained the complete phase diagram in the parameter space composed of the stimulus field, thermal noise, and network capacity. Our results show that the system improves its recognition ability for parallel stimulus. For orthogonal stimulus two recognition phases emerge with the system locking at the initialization or stimulated pattern. We confront our analytical results with numerical simulations for the noiseless case T=0 .

  18. Orthogonal design for scale invariant feature transform optimization

    NASA Astrophysics Data System (ADS)

    Ding, Xintao; Luo, Yonglong; Yi, Yunyun; Jie, Biao; Wang, Taochun; Bian, Weixin

    2016-09-01

    To improve object recognition capabilities in applications, we used orthogonal design (OD) to choose a group of optimal parameters in the parameter space of scale invariant feature transform (SIFT). In the case of global optimization (GOP) and local optimization (LOP) objectives, our aim is to show the operation of OD on the SIFT method. The GOP aims to increase the number of correctly detected true matches (NoCDTM) and the ratio of NoCDTM to all matches. In contrast, the LOP mainly aims to increase the performance of recall-precision. In detail, we first abstracted the SIFT method to a 9-way fixed-effect model with an interaction. Second, we designed a mixed orthogonal array, MA(64,23420,2), and its header table to optimize the SIFT parameters. Finally, two groups of parameters were obtained for GOP and LOP after orthogonal experiments and statistical analyses were implemented. Our experiments on four groups of data demonstrate that compared with the state-of-the-art methods, GOP can access more correct matches and is more effective against object recognition. In addition, LOP is favorable in terms of the recall-precision.

  19. Orthogonal dispersive spectral-domain optical coherence tomography.

    PubMed

    Bao, Wen; Ding, Zhihua; Li, Peng; Chen, Zhiyan; Shen, Yi; Wang, Chuan

    2014-04-21

    Ultrahigh depth range spectral domain optical coherence tomography (SDOCT) can be realized based on the orthogonal dispersive spectrometer consisted by a high spectral resolution virtually-imaged phased array (VIPA) and a low spectral resolution grating. However, two critical issues result in the challenge of obtaining desirable one-dimensional (1-D) spectra from the recorded two-dimensional (2-D) orthogonal spectra for high-quality OD-SDOCT imaging. One is the wavenumber mapping errors and the other is the periodic intensity modulations. The paper proposes a method for desirable reconstruction of 1-D spectra from the recorded 2-D orthogonal spectra. A sample etalon with identical parameters to the dispersive VIPA is used to determine the free spectrum range (FSR) of the VIPA, and spectral phases from two reflecting mirrors are further applied for broadband wavenumber calibration. The cascading of column spectra are performed from interval of four lines of column spectra, and four records of cascaded 1-D spectra are obtained and then averaged to alleviate the periodic intensity modulations. Broadband 1-D spectra are thus reconstructed with an ultrahigh spectral resolution. To demonstrate the feasibility of the proposed method, three typical samples are imaged by the OD-SDOCT system.

  20. Multilayer block copolymer meshes by orthogonal self-assembly

    PubMed Central

    Tavakkoli K. G., Amir; Nicaise, Samuel M.; Gadelrab, Karim R.; Alexander-Katz, Alfredo; Ross, Caroline A.; Berggren, Karl K.

    2016-01-01

    Continued scaling-down of lithographic-pattern feature sizes has brought templated self-assembly of block copolymers (BCPs) into the forefront of nanofabrication research. Technologies now exist that facilitate significant control over otherwise unorganized assembly of BCP microdomains to form both long-range and locally complex monolayer patterns. In contrast, the extension of this control into multilayers or 3D structures of BCP microdomains remains limited, despite the possible technological applications in next-generation devices. Here, we develop and analyse an orthogonal self-assembly method in which multiple layers of distinct-molecular-weight BCPs naturally produce nanomesh structures of cylindrical microdomains without requiring layer-by-layer alignment or high-resolution lithographic templating. The mechanisms for orthogonal self-assembly are investigated with both experiment and simulation, and we determine that the control over height and chemical preference of templates are critical process parameters. The method is employed to produce nanomeshes with the shapes of circles and Y-intersections, and is extended to produce three layers of orthogonally oriented cylinders. PMID:26796218

  1. Modular and orthogonal synthesis of hybrid polymers and networks.

    PubMed

    Liu, Shuang; Dicker, Kevin T; Jia, Xinqiao

    2015-03-28

    Biomaterials scientists strive to develop polymeric materials with distinct chemical make-up, complex molecular architectures, robust mechanical properties and defined biological functions by drawing inspirations from biological systems. Salient features of biological designs include (1) repetitive presentation of basic motifs; and (2) efficient integration of diverse building blocks. Thus, an appealing approach to biomaterials synthesis is to combine synthetic and natural building blocks in a modular fashion employing novel chemical methods. Over the past decade, orthogonal chemistries have become powerful enabling tools for the modular synthesis of advanced biomaterials. These reactions require building blocks with complementary functionalities, occur under mild conditions in the presence of biological molecules and living cells and proceed with high yield and exceptional selectivity. These chemistries have facilitated the construction of complex polymers and networks in a step-growth fashion, allowing facile modulation of materials properties by simple variations of the building blocks. In this review, we first summarize features of several types of orthogonal chemistries. We then discuss recent progress in the synthesis of step growth linear polymers, dendrimers and networks that find application in drug delivery, 3D cell culture and tissue engineering. Overall, orthogonal reactions and modulular synthesis have not only minimized the steps needed for the desired chemical transformations but also maximized the diversity and functionality of the final products. The modular nature of the design, combined with the potential synergistic effect of the hybrid system, will likely result in novel hydrogel matrices with robust structures and defined functions.

  2. Limited-memory adaptive snapshot selection for proper orthogonal decomposition

    SciTech Connect

    Oxberry, Geoffrey M.; Kostova-Vassilevska, Tanya; Arrighi, Bill; Chand, Kyle

    2015-04-02

    Reduced order models are useful for accelerating simulations in many-query contexts, such as optimization, uncertainty quantification, and sensitivity analysis. However, offline training of reduced order models can have prohibitively expensive memory and floating-point operation costs in high-performance computing applications, where memory per core is limited. To overcome this limitation for proper orthogonal decomposition, we propose a novel adaptive selection method for snapshots in time that limits offline training costs by selecting snapshots according an error control mechanism similar to that found in adaptive time-stepping ordinary differential equation solvers. The error estimator used in this work is related to theory bounding the approximation error in time of proper orthogonal decomposition-based reduced order models, and memory usage is minimized by computing the singular value decomposition using a single-pass incremental algorithm. Results for a viscous Burgers’ test problem demonstrate convergence in the limit as the algorithm error tolerances go to zero; in this limit, the full order model is recovered to within discretization error. The resulting method can be used on supercomputers to generate proper orthogonal decomposition-based reduced order models, or as a subroutine within hyperreduction algorithms that require taking snapshots in time, or within greedy algorithms for sampling parameter space.

  3. Phosphorylation of Human CTP Synthetase 1 by Protein Kinase A: IDENTIFICATION OF Thr455 AS A MAJOR SITE OF PHOSPHORYLATION*

    PubMed Central

    Choi, Mal-Gi; Carman, George M.

    2007-01-01

    CTP synthetase is an essential enzyme that generates the CTP required for the synthesis of nucleic acids and membrane phospholipids. In this work, we examined the phosphorylation of the human CTPS1-encoded CTP synthetase 1 by protein kinase A. CTP synthetase 1 was expressed and purified from a Saccharomyces cerevisiae ura7Δ ura8Δ double mutant that lacks CTP synthetase activity. Using purified CTP synthetase 1 as a substrate, protein kinase A activity was time- and dose-dependent. The phosphorylation, which primarily occurred on a threonine residue, was accompanied by a 50% decrease in CTP synthetase 1 activity. The synthetic peptide LGKRRTLFQT that contains the protein kinase A motif for Thr455 was a substrate for protein kinase A. A Thr455 to Ala (T455A) mutation in CTP synthetase 1 was constructed by site-directed mutagenesis and was expressed and purified from the S. cerevisiae ura7Δ ura8Δ mutant. The T455A mutation caused a 78% decrease in protein kinase A phosphorylation, and the loss of the phosphothreonine residue and a major phosphopeptide that were present in the purified wild type enzyme phosphorylated by protein kinase A. The CTP synthetase 1 activity of the T455A mutant enzyme was 2-fold higher than the wild type enzyme. In addition, the T455A mutation caused a 44% decrease in the amount of human CTP synthetase 1 that was phosphorylated in S. cerevisiae cells, and this was accompanied by a 2.5-fold increase in the cellular concentration of CTP and a 1.5-fold increase in the choline-dependent synthesis of phosphatidylcholine. PMID:17189248

  4. The effect of glial glutamine synthetase inhibition on recognition and temporal memories in the rat.

    PubMed

    Kant, Deepika; Tripathi, Shweta; Qureshi, Munazah F; Tripathi, Shweta; Pandey, Swati; Singh, Gunjan; Kumar, Tankesh; Mir, Fayaz A; Jha, Sushil K

    2014-02-07

    The glutamate neurotransmitter is intrinsically involved in learning and memory. Glial glutamine synthetase enzyme synthesizes glutamine, which helps maintain the optimal neuronal glutamate level. However, the role of glutamine synthetase in learning and memory remains unclear. Using associative trace learning task, we investigated the effects of methionine sulfoximine (MSO) (glutamine synthetase inhibitor) on recognition and temporal memories. MSO and vehicle were injected (i.p.) three hours before training in separate groups of male Wistar rats (n=11). Animals were trained to obtain fruit juice after following a set of sequential events. Initially, house-light was presented for 15s followed by 5s trace interval. Thereafter, juice was given for 20s followed by 20s inter-presentation interval. A total of 75 presentations were made over five sessions during the training and testing periods. The average number of head entries to obtain juice per session and during individual phases at different time intervals was accounted as an outcome measure of recognition and temporal memories. The total head entries in MSO and vehicle treated animals were comparable on training and testing days. However, it was 174.90% (p=0.08), 270.61% (p<0.05), 143.20% (p<0.05) more on training day and 270.33% (p<0.05), 157.94% (p<0.05), 170.42% (p<0.05) more on testing day, during the house-light, trace-interval and inter-presentation interval phases in MSO animals. Glutamine synthetase inhibition did not induce recognition memory deficit, while temporal memory was altered, suggesting that glutamine synthetase modulates some aspects of mnemonic processes.

  5. Molecular cloning and characterization of glutamine synthetase, a tegumental protein from Schistosoma japonicum.

    PubMed

    Qiu, Chunhui; Hong, Yang; Cao, Yan; Wang, Fei; Fu, Zhiqiang; Shi, Yaojun; Wei, Meimei; Liu, Shengfa; Lin, Jiaojiao

    2012-12-01

    Glutamine synthetase catalyzes the synthesis of glutamine, providing nitrogen for the production of purines, pyrimidines, amino acids, and other compounds required in many pivotal cellular events. Herein, a full-length cDNA encoding Schistosoma japonicum glutamine synthetase (SjGS) was isolated from 21-day schistosomes. The entire open reading frame of SjGS contains a 1,095-bp coding region corresponding to 364 amino acids with a calculated molecular weight of 40.7 kDa. NCBIP blast shows that the putative amino acid of SjGS contains a classic β-grasp domain and a catalytic domain of glutamine synthetase. The relative mRNA expression of SjGS was evaluated in 7-, 13-, 21-, 28-, 35-, and 42-day worms of S. japonicum in the final host and higher expression at day 21, and 42 worms were observed. This protein was also detected in worm extracts using Western blot. Immunofluorescence studies indicated that the SjGS protein was mainly distributed on tegument and parenchyma in 28-day adult worms. The recombinant glutamine synthetase with a molecular weight of 45 kDa was expressed in Escherichia coli and purified in its active form. The enzyme activity of the recombinant protein was 3.30 ± 0.67 U.μg-1. The enzyme activity was highly stable over a wide range of pH (6-9) and temperature (25-40 °C) under physiological conditions. The transcription of SjGS was upregulated in praziquantel-treated worms at 2-, 4-, and 24-h posttreatment compared with the untreated control. As a first step towards the clarification of the role of glutamine synthetase in schistosome species, we have cloned and characterized cDNAs encoding SjGS in S. japonicum, and the data presented suggest that SjGS is an important molecule in the development of the schistosome.

  6. Regulation of the intersubunit ammonia tunnel in Mycobacterium tuberculosis glutamine-dependent NAD[superscript +] synthetase

    SciTech Connect

    Chuenchor, Watchalee; Doukov, Tzanko I.; Resto, Melissa; Chang, Andrew; Gerratana, Barbara

    2012-08-31

    Glutamine-dependent NAD{sup +} synthetase is an essential enzyme and a validated drug target in Mycobacterium tuberculosis (mtuNadE). It catalyses the ATP-dependent formation of NAD{sup +} from NaAD{sup +} (nicotinic acid-adenine dinucleotide) at the synthetase active site and glutamine hydrolysis at the glutaminase active site. An ammonia tunnel 40 {angstrom} (1 {angstrom} = 0.1 nm) long allows transfer of ammonia from one active site to the other. The enzyme displays stringent kinetic synergism; however, its regulatory mechanism is unclear. In the present paper, we report the structures of the inactive glutaminase C176A variant in an apo form and in three synthetase-ligand complexes with substrates (NaAD{sup +}/ATP), substrate analogue {l_brace}NaAD{sup +}/AMP-CPP (adenosine 5'-[{alpha},{beta}-methylene]triphosphate){r_brace} and intermediate analogues (NaAD{sup +}/AMP/PPi), as well as the structure of wild-type mtuNadE in a product complex (NAD{sup +}/AMP/PPi/glutamate). This series of structures provides snapshots of the ammonia tunnel during the catalytic cycle supported also by kinetics and mutagenesis studies. Three major constriction sites are observed in the tunnel: (i) at the entrance near the glutaminase active site; (ii) in the middle of the tunnel; and (iii) at the end near the synthetase active site. Variation in the number and radius of the tunnel constrictions is apparent in the crystal structures and is related to ligand binding at the synthetase domain. These results provide new insight into the regulation of ammonia transport in the intermolecular tunnel of mtuNadE.

  7. Structural analysis of the active site geometry of N5-carboxyaminoimidazole ribonucleotide synthetase from Escherichia coli.

    PubMed

    Thoden, James B; Holden, Hazel M; Firestine, Steven M

    2008-12-16

    N(5)-Carboxyaminoimidazole ribonucleotide synthetase (N(5)-CAIR synthetase) converts 5-aminoimidazole ribonucleotide (AIR), MgATP, and bicarbonate into N(5)-CAIR, MgADP, and P(i). The enzyme is required for de novo purine biosynthesis in microbes yet is not found in humans suggesting that it represents an ideal and unexplored target for antimicrobial drug design. Here we report the X-ray structures of N(5)-CAIR synthetase from Escherichia coli with either MgATP or MgADP/P(i) bound in the active site cleft. These structures, determined to 1.6-A resolution, provide detailed information regarding the active site geometry before and after ATP hydrolysis. In both structures, two magnesium ions are observed. Each of these is octahedrally coordinated, and the carboxylate side chain of Glu238 bridges them. For the structure of the MgADP/P(i) complex, crystals were grown in the presence of AIR and MgATP. No electron density was observed for AIR, and the electron density corresponding to the nucleotide clearly revealed the presence of ADP and P(i) rather than ATP. The bound P(i) shifts by approximately 3 A relative to the gamma-phosphoryl group of ATP and forms electrostatic interactions with the side chains of Arg242 and His244. Since the reaction mechanism of N(5)-CAIR synthetase is believed to proceed via a carboxyphosphate intermediate, we propose that the location of the inorganic phosphate represents the binding site for stabilization of this reactive species. Using the information derived from the two structures reported here, coupled with molecular modeling, we propose a catalytic mechanism for N(5)-CAIR synthetase.

  8. A genomically modified Escherichia coli strain carrying an orthogonal E. coli histidyl-tRNA synthetase•tRNA(His) pair.

    PubMed

    Englert, Markus; Vargas-Rodriguez, Oscar; Reynolds, Noah M; Wang, Yane-Shih; Söll, Dieter; Umehara, Takuya

    2017-03-10

    Development of new aminoacyl-tRNA synthetase (aaRS)•tRNA pairs is central for incorporation of novel non-canonical amino acids (ncAAs) into proteins via genetic code expansion (GCE). The Escherichia coli and Caulobacter crescentus histidyl-tRNA synthetases (HisRS) evolved divergent mechanisms of tRNA(His) recognition that prevent their cross-reactivity. Although the E. coli HisRS•tRNA(His) pair is a good candidate for GCE, its use in C. crescentus is limited by the lack of established genetic selection methods and by the low transformation efficiency of C. crescentus. E. coli was genetically engineered to use a C. crescentus HisRS•tRNA(His) pair. Super-folder green fluorescent protein (sfGFP) and chloramphenicol acetyltransferase (CAT) were used as reporters for read-through assays. A library of 313 ncAAs coupled with the sfGFP reporter system was employed to investigate the specificity of E. coli HisRS in vivo. A genomically modified E. coli strain (named MEOV1) was created. MEVO1 requires an active C. crescentus HisRS•tRNA(His) pair for growth, and displays a similar doubling time as the parental E. coli strain. sfGFP- and CAT-based assays showed that the E. coli HisRS•tRNA(His) pair is orthogonal in MEOV1 cells. A mutation in the anticodon loop of E. coli tRNA(His)CUA elevated its suppression efficiency by 2-fold. The C. crescentus HisRS•tRNA(His) pair functionally complements an E. coli ΔhisS strain. The E. coli HisRS•tRNA(His) is orthogonal in MEOV1 cells. E. coli tRNA(His)CUA is an efficient amber suppressor in MEOV1. We developed a platform that allows protein engineering of E. coli HisRS that should facilitate GCE in E. coli. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Hydroxamate-based colorimetric assay to assess amide bond formation by adenylation domain of nonribosomal peptide synthetases.

    PubMed

    Hara, Ryotaro; Suzuki, Ryohei; Kino, Kuniki

    2015-05-15

    We demonstrated the usefulness of a hydroxamate-based colorimetric assay for predicting amide bond formation (through an aminoacyl-AMP intermediate) by the adenylation domain of nonribosomal peptide synthetases. By using a typical adenylation domain of tyrocidine synthetase (involved in tyrocidine biosynthesis), we confirmed the correlation between the absorbance at 490 nm of the l-Trp-hydroxamate-Fe(3+) complex and the formation of l-Trp-l-Pro, where l-Pro was used instead of hydroxylamine. Furthermore, this assay was adapted to the adenylation domains of surfactin synthetase (involved in surfactin biosynthesis) and bacitracin synthetase (involved in bacitracin biosynthesis). Consequently, the formation of various aminoacyl l-Pro formations was observed.

  10. Molecular cloning and regulation of expression of the genes for initiation factor 3 and two aminoacyl-tRNA synthetases.

    PubMed Central

    Elseviers, D; Gallagher, P; Hoffman, A; Weinberg, B; Schwartz, I

    1982-01-01

    A 22-kilobase fragment of the Escherichia coli chromosome which contains the genes for translation initiation factor 3, phenylalanyl-tRNA synthetase, and threonyl-tRNA synthetase was cloned into plasmid pACYC184. The hybrid plasmid (designated pID1) complements a temperature-sensitive pheS lesion in E. coli NP37. pID1-transformed NP37 overproduce initiation factor 3 and phenylalanyl-tRNA synthetase. Gene expression from pID1 was studied in vitro in a coupled transcription-translation system and in minicells. The results suggest that the genes for initiation factor 3 and phenylalanyl- and threonyl-tRNA synthetase are regulated by different mechanisms. Images PMID:6749810

  11. Molecular cloning and regulation of expression of the genes for initiation factor 3 and two aminoacyl-tRNA synthetases.

    PubMed

    Elseviers, D; Gallagher, P; Hoffman, A; Weinberg, B; Schwartz, I

    1982-10-01

    A 22-kilobase fragment of the Escherichia coli chromosome which contains the genes for translation initiation factor 3, phenylalanyl-tRNA synthetase, and threonyl-tRNA synthetase was cloned into plasmid pACYC184. The hybrid plasmid (designated pID1) complements a temperature-sensitive pheS lesion in E. coli NP37. pID1-transformed NP37 overproduce initiation factor 3 and phenylalanyl-tRNA synthetase. Gene expression from pID1 was studied in vitro in a coupled transcription-translation system and in minicells. The results suggest that the genes for initiation factor 3 and phenylalanyl- and threonyl-tRNA synthetase are regulated by different mechanisms.

  12. Integrated, Dual Orthogonal Antennas for Polarimetric Ground Penetrating Radar

    NASA Astrophysics Data System (ADS)

    Pauli, Mario; Wiesbeck, Werner

    2015-04-01

    Ground penetrating radar systems are mostly equipped with single polarized antennas, for example with single linear polarization or with circular polarization. The radiated waves are partly reflected at the ground surface and very often the penetrating waves are distorted in their polarization. The distortion depends on the ground homogeneity and the orientation of the antennas relative to the ground structure. The received signals from the reflecting objects may most times only be classified according to their coverage and intensity. This makes the recognition of the objects difficult or impossible. In airborne and spaceborne Remote Sensing the systems are meanwhile mostly equipped with front ends with dual orthogonal polarized antennas for a full polarimetric operation. The received signals, registered in 2x2 scattering matrices according to co- and cross polarization, are processed for the evaluation of all features of the targets. Ground penetrating radars could also profit from the scientific results of Remote Sensing. The classification of detected objects for their structure and orientation requires more information in the reflected signal than can be measured with a single polarization [1, 2]. In this paper dual linear, orthogonal polarized antennas with a common single, frequency independent phase center, are presented [3]. The relative bandwidth of these antennas can be 1:3, up to 1:4. The antenna is designed to work in the frequency range between 3 GHz and 11 GHz, but can be easily adapted to the GPR frequency range by scaling. The size of the antenna scaled for operation in typical GPR frequencies would approximately be 20 by 20 cm2. By the implementation in a dielectric carrier it could be reduced in size if required. The major problem for ultra wide band, dual polarized antennas is the frequency independent feed network, realizing the required phase shifts. For these antennas a network, which is frequency independent over a wide range, has been

  13. Resolution enhancement of computed radiography images using two orthogonal tilts

    NASA Astrophysics Data System (ADS)

    Pollmann, Steven I.; Norley, Chris J. D.; Yuan, Xunhua; Holdsworth, David W.

    2012-03-01

    Limitations to the spatial resolution of current digital x-ray systems are bounded by the physical characteristics of the xray detector. However, the need to image smaller structures provides motivation to develop high-resolution x-ray detector systems for use with computed radiographic, and tomographic x-ray systems. We report the implementation of a tilted detector technique (TDT) to attain near isotropic resolution enhancement by combining two orthogonal image views, acquired with existing detector hardware tilted at a fixed angle. Images were acquired using a ceiling-mounted x-ray unit (Proteus XR/a, GE Medical Systems, 50kVp, 250mAs). Images were digitized using a Fujifilm Capsula X CR system, from a 35×43cm detector cassette placed on an angulated stand, featuring a 3520×4280 image matrix with an in-plane pixel spacing of 0.1mm. Three images were acquired: two for use with our TDT; and one for comparison, with no detector tilt. Performance was determined by using two line-pair phantoms (Models 07-521 and 07-533, Nuclear Associates) placed orthogonally to each other in the field of view. Custom software corrected for perspective distortion, co-registered and combined the tilted-detector images into a single higher-resolution image. Following unwarping and co-registration, the limiting spatial resolution of an image obtained via the weighted combination of the two orthogonal views (8 lp/mm) is found to be superior to that of a single view acquired with no detector tilt (5 lp/mm). This novel technique shows significant improvement in the spatial resolution of x-ray image acquisitions, using existing x-ray components and detector hardware.

  14. Collinearity and orthogonality of endmembers in linear spectral unmixing

    NASA Astrophysics Data System (ADS)

    Van der Meer, Freek D.; Jia, Xiuping

    2012-08-01

    Contrary to image classification, spectral unmixing techniques allow to derive abundance/fractional cover estimates for selected endmembers within the volume of a pixel. Mathematically the solution to the mixing problem is resolving a set of linear equations using least squares approaches. Practically this is done using singular value deconvolution of the endmember matrix inversion. This solution assumes orthogonality of the endmembers which determines the orthogonality of the matrix. If endmembers are highly correlated (thus collinearity or multi-collinearity occurs), the matrix becomes non-orthogonal, the inversion unstable and the inverse or estimated fractions highly sensitive to random error (e.g., noise). In practice, collinearity almost always exists but it is typically overlooked or ignored, hence with this overview we wish to create awareness to the issue and offer approaches to deal with the problem. The first part of the paper highlights the problem using a numerical example. It is shown how collinearity amplifies the error in the endmember matrix inversion. In the next paragraph we propose measures to quantify the level of (multi)collinearity in the endmember matrix: a weighted multiple correlation measure, the variance inflation factor, the partial regression coefficient. The remainder of the paper is dedicated to approaches to mitigate the problem: excluding endmembers, decorrelating endmembers, iterative approaches for endmember selection and we propose an adjustment to the unmixing equation which could be further explored. In conclusion, collinearity hampers the use of fractional abundance estimates. There is no single recipe to successfully combat this problem but in all mixture models collinearity should be tested and avoided as much as possible.

  15. Adaptive PID control based on orthogonal endocrine neural networks.

    PubMed

    Milovanović, Miroslav B; Antić, Dragan S; Milojković, Marko T; Nikolić, Saša S; Perić, Staniša Lj; Spasić, Miodrag D

    2016-12-01

    A new intelligent hybrid structure used for online tuning of a PID controller is proposed in this paper. The structure is based on two adaptive neural networks, both with built-in Chebyshev orthogonal polynomials. First substructure network is a regular orthogonal neural network with implemented artificial endocrine factor (OENN), in the form of environmental stimuli, to its weights. It is used for approximation of control signals and for processing system deviation/disturbance signals which are introduced in the form of environmental stimuli. The output values of OENN are used to calculate artificial environmental stimuli (AES), which represent required adaptation measure of a second network-orthogonal endocrine adaptive neuro-fuzzy inference system (OEANFIS). OEANFIS is used to process control, output and error signals of a system and to generate adjustable values of proportional, derivative, and integral parameters, used for online tuning of a PID controller. The developed structure is experimentally tested on a laboratory model of the 3D crane system in terms of analysing tracking performances and deviation signals (error signals) of a payload. OENN-OEANFIS performances are compared with traditional PID and 6 intelligent PID type controllers. Tracking performance comparisons (in transient and steady-state period) showed that the proposed adaptive controller possesses performances within the range of other tested controllers. The main contribution of OENN-OEANFIS structure is significant minimization of deviation signals (17%-79%) compared to other controllers. It is recommended to exploit it when dealing with a highly nonlinear system which operates in the presence of undesirable disturbances.

  16. Effect of single base substitutions at glycine-870 codon of gramicidin S synthetase 2 gene on proline activation.

    PubMed

    Tokita, K; Hori, K; Kurotsu, T; Kanda, M; Saito, Y

    1993-10-01

    The mutant gene coding for a proline-activating domain (grs2-pro) was cloned and sequenced from Bacillus brevis Nagano, BII-3 strain, which produces gramicidin S synthetase 2 defective in proline-activation. By comparison of the nucleotide sequence with the wild-type sequence, a single point mutation was found at the 2609th guanine, which was replaced with adenine, resulting in the change of the 870th glycine to glutamic acid. Homology search for the deduced amino acid sequence of grs2-pro gene revealed that the 870th glycine was conserved in adenylate-forming enzymes, and its flanking sequence was highly conserved among the aminoacyl adenylate-forming enzymes, such as antibiotic peptide synthetases: gramicidin S synthetase 1 and 2 (GS1, GS2), tyrocidine synthetase 1 (TS1), and delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase (ACVS); and other aminoacyl adenylation enzymes: alpha-aminoadipate reductase (LYS2), EntF, and AngR. On the other hand, this flanking sequence was not conserved in the other adenylate-forming enzymes lacking amino acid activation, such as acetyl-CoA synthetase, long-chain acyl-CoA synthetase, luciferase, and 4-coumarate CoA ligase. Single base substitutions at the 870th GGG codon were carried out by oligonucleotide site-directed mutagenesis. Four mutagenized clones were isolated, containing grs2-pro genes which exchange 870-Gly for alanine, valine, arginine, and tryptophan. The translated products from these clones could scarcely catalyze proline-dependent ATP-32PPi exchange reaction. The coil structure of 870-Gly region was lost in the mutants. These results suggest that the 870-Gly residue of grs2-pro protein is essential for aminoacyl-adenylation in the antibiotic peptide synthetase family.

  17. Constrained reduced-order models based on proper orthogonal decomposition

    DOE PAGES

    Reddy, Sohail R.; Freno, Brian Andrew; Cizmas, Paul G. A.; ...

    2017-04-09

    A novel approach is presented to constrain reduced-order models (ROM) based on proper orthogonal decomposition (POD). The Karush–Kuhn–Tucker (KKT) conditions were applied to the traditional reduced-order model to constrain the solution to user-defined bounds. The constrained reduced-order model (C-ROM) was applied and validated against the analytical solution to the first-order wave equation. C-ROM was also applied to the analysis of fluidized beds. Lastly, it was shown that the ROM and C-ROM produced accurate results and that C-ROM was less sensitive to error propagation through time than the ROM.

  18. Orthogonal (transverse) arrangements of actin in endothelia and fibroblasts

    PubMed Central

    Curtis, Adam; Aitchison, Gregor; Tsapikouni, Theodora

    2006-01-01

    Though actin filaments running across the cell (transverse actin) have been occasionally reported for epithelial cells in groups and for cells growing on fibres, there has been no report heretofore of transverse actin in cells grown on planar substrata. This paper describes evidence in support of this possibility derived from actin staining, polarization microscopy and force measurements. The paper introduces two new methods for detecting the orientation and activity of contractile elements in cells. The orthogonal actin is most obvious in cells grown on groove ridge structures, but can be detected in cells grown on flat surfaces. PMID:17015307

  19. Capillary-Driven Flow in Liquid Filaments Connecting Orthogonal Channels

    NASA Technical Reports Server (NTRS)

    Allen, Jeffrey S.

    2005-01-01

    Capillary phenomena plays an important role in the management of product water in PEM fuel cells because of the length scales associated with the porous layers and the gas flow channels. The distribution of liquid water within the network of gas flow channels can be dramatically altered by capillary flow. We experimentally demonstrate the rapid movement of significant volumes of liquid via capillarity through thin liquid films which connect orthogonal channels. The microfluidic experiments discussed provide a good benchmark against which the proper modeling of capillarity by computational models may be tested. The effect of surface wettability, as expressed through the contact angle, on capillary flow will also be discussed.

  20. Photoacoustic tomography using orthogonal Fabry-Pérot sensors

    NASA Astrophysics Data System (ADS)

    Ellwood, Robert; Ogunlade, Olumide; Zhang, Edward; Beard, Paul; Cox, Ben

    2017-04-01

    Fabry-Pérot sensors have been used to produce in-vivo photoacoustic images of exquisite quality. However, for ease of construction and interrogation, they are produced in a planar form. Planar arrays suffer from a limited detection aperture, which leads to artifacts in the reconstruction of the initial pressure distribution. Here, an L-shaped detection geometry is described that allows a greater field of view by placing a second planar array orthogonal to the first. This captures data from the deeper lying regions of interest and mitigates the limited view, thus reducing artifacts in the reconstructed initial pressure distribution.

  1. Quantum memory in an orthogonal geometry of silenced echo retrieval

    NASA Astrophysics Data System (ADS)

    Gerasimov, K. I.; Minnegaliev, M. M.; Moiseev, S. A.; Urmancheev, R. V.; Chanelière, T.; Louchet-Chauvet, A.

    2017-08-01

    We experimentally realize a quantum-memory protocol based on retrieval of silenced echo (ROSE) in Tm3+:Y3Al5O12 crystal in an orthogonal geometry of the signal and control light fields. The silenced echo signal revival efficiency of 13% with 36 μs storage time is demonstrated. To achieve that we implemented a high-precision atomic coherence control via amplitude- and phase-modulated laser pulses. We also discuss capabilities of this configuration, ways to increase quantum efficiency and to combine it with a single-mode optical cavity.

  2. Magnetic particle detection in unshielded environment using orthogonal fluxgate gradiometer

    SciTech Connect

    Elrefai, Ahmed L. Sasada, Ichiro

    2015-05-07

    A new detection system for magnetic particles, which can operate in an unshielded environment, is developed using a fundamental mode orthogonal fluxgate gradiometer. The proposed detection system offers the advantages of cost, size, and weight reduction as compared to contamination detection systems using superconducting quantum interference device sensor. The detection system can be used to detect metallic contamination in foods or lithium ion battery production lines. The system has been investigated numerically to optimize various design parameters of the system. Experimental setup has been developed to evaluate some of the numerically predicted results. Steel balls were successfully detected down to the diameter of 50 μm.

  3. Conditioning Analysis of Incomplete Cholesky Factorizations with Orthogonal Dropping

    SciTech Connect

    Napov, Artem

    2013-08-01

    The analysis of preconditioners based on incomplete Cholesky factorization in which the neglected (dropped) components are orthogonal to the approximations being kept is presented. General estimate for the condition number of the preconditioned system is given which only depends on the accuracy of individual approximations. The estimate is further improved if, for instance, only the newly computed rows of the factor are modified during each approximation step. In this latter case it is further shown to be sharp. The analysis is illustrated with some existing factorizations in the context of discretized elliptic partial differential equations.

  4. Some properties of multiple orthogonal polynomials associated with Macdonald functions

    NASA Astrophysics Data System (ADS)

    Coussement, Els; van Assche, Walter

    2001-08-01

    Multiple orthogonal polynomials corresponding to two weights on [0,[infinity]) associated with modified Bessel functions (Macdonald functions) K[nu] and K[nu]+1 were introduced in Van Assche, Yakubovich (Integral Transforms Special Funct. 9 (2000) 229-244) and recently also studied by Ben Cheikh, Douak (Meth. Appl. Anal., to appear). We obtain explicit formulas for type I vector polynomials (An,n,Bn,n) and (An+1,n,Bn+1,n) and for type II polynomials Pn,n and Pn+1,n. We also obtain generating functions for types I and II polynomials.

  5. Variability in GRB light curves: Introducing Orthogonal Matching Pursuit

    NASA Astrophysics Data System (ADS)

    Dereli, Husne; Bégué, Damien; Ryde, Felix

    2016-07-01

    Constraining the variability of GRBs is important as it is one of the few keys to estimate many unknown parameters, such as the emission radius, the Lorentz factor, the size of the progenitor. In this work, we introduced the Orthogonal Matching Pursuit (OMP) method to study GRB light curves and to compute the minimum time variability of GRBs. Commonly used in medical sciences, this method reconstructs a signal by choosing among predefined functional shapes. We will discuss the implementation of the code, and compare its performances with those of other dedicated methods (Haar wavelet analysis, peak finding algorithm and step wise filter correlation).

  6. Generation, modulation and reception of orthogonally polarized signals

    NASA Technical Reports Server (NTRS)

    Lesh, J. R.

    1985-01-01

    The radiating electromagnetic field of a dipole antenna is considered. By allowing the dipole to rotate about its midpoint, one can construct an entire set of signal polarizations wherein distinct members of the set are mutually orthogonal. It is shown how these signals can be modulated and demodulated to convey information. These ideas are then generalized, and both balanced, as well as unbalanced quadrapole modulations are defined. Methods of receiver synchronization to such signals are described, as well as their potential application to multiple access and anti-jam communications.

  7. Dendron avidity platforms with orthogonal focal point coupling site

    NASA Astrophysics Data System (ADS)

    McNerny, Daniel Quinn

    This thesis explores the design and synthesis of bifunctional or modular platforms from poly(amidoamine) (PAMAM) dendrons. PAMAM dendrons with an orthogonal focal point are evaluated, testing several click chemistry reactions for high conversion and mild conditions. The orthogonal reaction chemistry used at the dendron focal point gives a precise 1:1 ratio of the attachment of multiple functionalities to a small molecular weight, chemically stable high avidity molecule. In the first component of the thesis, dendrons were synthesized with c(RGDyK) peptide on the surface to create a scaffold for cellular targeting and multivalent binding. Binary dendron-RGD conjugates were synthesized with a single imaging agent, therapeutic drug, or additional functionalized dendron at the focal point after a copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC) click reaction. The targeted-dendron platform was shown to specifically target alphaVbeta3 integrin expressing human umbilical vein endothelial cells (HUVEC) and human glioblastoma cells (U87MG) in vitro via flow cytometry. Specific targeting of the dendron-RGD platform was further confirmed by confocal microscopy. Biological activity of the targeted drug conjugate was confirmed via XTT assay. The remainder of the thesis explores click chemistry reactions that do not require a metal catalyst, which may cause undesired toxicity for some biological applications. Thiol-based click chemistry, specifically the thiol-ene and thiol-yne reactions, is explored on dendron platforms. The thiol click reactions provide an improved efficiency, compared to CuAAC, by reaching quantitative conversion of the focal point in most cases. The thiol click reactions suffer from some setbacks: the need for a thermal or photoinitiator may prevent the conjugation of some functional ligands and the thiol chemistry is more prone to side reactions. Finally, strain-promoted alkyne-azide cycloadditions are examined. The ring-strain click chemistry

  8. Evaluation of pairwise distances among orthogonal grid points in hypercube

    NASA Astrophysics Data System (ADS)

    Sadílek, Václav; Vořechovský, Miroslav; Šmídová, Magdalena

    2017-07-01

    This article describes an effective evaluation of pairwise distances among points that form a regular orthogonal grid inside a unit hypercube. Such an regular arrangement of points have many applications one of which is the Design of Experiments - the arrangement is known as the Full Factorial Design. The proposed process of calculation exploits the repeated point projections. Using combinatorial rules, the paper presents a closed-form exact formula that enables faster evaluation of pairwise distances and their counts compared to naive approach that enumerates the list of all possible pairs of points.

  9. Orthogonality between scales in a renormalization group for fermions

    NASA Astrophysics Data System (ADS)

    Pereira, Emmanuel

    1995-02-01

    Having in mind the development of a technical tool to treat fermionic systems, we propose a Kadanoff-Wilson block renormalization transformation employing unusual averages (an inevitable artifact due to the specificity of lattice fermions and to the desired transformation properties). The free propagator is decomposed into operators associated to different momentum scales and with orthogonal relations, and the effective actions generated from the Dirac operator by the transformations present uniform exponential decay. We argue to show the usefulness of the formalism to study correlation functions of interacting fermions.

  10. Photoacoustic tomography using orthogonal Fabry-Pérot sensors.

    PubMed

    Ellwood, Robert; Ogunlade, Olumide; Zhang, Edward; Beard, Paul; Cox, Ben

    2017-04-01

    Fabry–Pérot sensors have been used to produce in-vivo photoacoustic images of exquisite quality. However, for ease of construction and interrogation, they are produced in a planar form. Planar arrays suffer from a limited detection aperture, which leads to artifacts in the reconstruction of the initial pressure distribution. Here, an L-shaped detection geometry is described that allows a greater field of view by placing a second planar array orthogonal to the first. This captures data from the deeper lying regions of interest and mitigates the limited view, thus reducing artifacts in the reconstructed initial pressure distribution.

  11. Structural performance of orthogonal tetrahedral truss Space-Station configurations

    NASA Technical Reports Server (NTRS)

    Dorsey, J. T.

    1984-01-01

    Two 150 kW space station configurations constructed with the orthogonal tetrahedral truss concept are described. One space station consists of a large central platform and two rotating solar wing arrays and the other consists of a long central keel with two rotating arrays. The dynamic characteristics of each configuration are obtained with and without nonstructural components present. The variation in frequencies and mass moments of inertia due to rotation of the two solar wing arrays are given for the long keel space station configuration. The structural performance of the solar wing array is assessed for cases where individual critical struts fail in the array support truss.

  12. Optimal approximation of harmonic growth clusters by orthogonal polynomials

    SciTech Connect

    Teodorescu, Razvan

    2008-01-01

    Interface dynamics in two-dimensional systems with a maximal number of conservation laws gives an accurate theoreticaI model for many physical processes, from the hydrodynamics of immiscible, viscous flows (zero surface-tension limit of Hele-Shaw flows), to the granular dynamics of hard spheres, and even diffusion-limited aggregation. Although a complete solution for the continuum case exists, efficient approximations of the boundary evolution are very useful due to their practical applications. In this article, the approximation scheme based on orthogonal polynomials with a deformed Gaussian kernel is discussed, as well as relations to potential theory.

  13. Analysis of secured Optical Orthogonal Frequency Division Multiplexed System

    NASA Astrophysics Data System (ADS)

    Gill, Harsimranjit Singh; Bhatia, Kamaljit Singh; Gill, Sandeep Singh

    2017-05-01

    In this paper, security issues for optical orthogonal frequency division multiplexed (OFDM) systems are emphasized. The encryption has been done on the data of coded OFDM symbols using data encryption standard (DES) algorithm before transmitting through the fiber. The results obtained justify that the DES provides better security to the input data without further bandwidth requirement. The data is transmitted to a distance of 1,000 km in a single-mode fiber with 16-quadrature amplitude modulation. The peak-to-average power ratio and optical signal-to-noise ratio of secure coded OFDM signal is fairly better than the conventional OFDM signal.

  14. Orthogonal frequency division multiplexing simulation based on MATLAB

    NASA Astrophysics Data System (ADS)

    Qiao, Yuan

    2017-09-01

    OFDM (Orthogonal Frequency Division Multiplexing) is one of the core technologies in the fourth generation mobile communication system. It is a widely-used method of the multi-carrier modulations based on IFFT and FFT transform, it can achieve the lowest complexity and effectively combat frequency selective fading. In this paper, we successfully use MATLAB to do the simulation of OFDM, and obtained good results, in which successful recovery out of the original signal under real channel condition, and error is less than 5% with the original signal.

  15. Magnetic particle detection in unshielded environment using orthogonal fluxgate gradiometer

    NASA Astrophysics Data System (ADS)

    Elrefai, Ahmed L.; Sasada, Ichiro

    2015-05-01

    A new detection system for magnetic particles, which can operate in an unshielded environment, is developed using a fundamental mode orthogonal fluxgate gradiometer. The proposed detection system offers the advantages of cost, size, and weight reduction as compared to contamination detection systems using superconducting quantum interference device sensor. The detection system can be used to detect metallic contamination in foods or lithium ion battery production lines. The system has been investigated numerically to optimize various design parameters of the system. Experimental setup has been developed to evaluate some of the numerically predicted results. Steel balls were successfully detected down to the diameter of 50 μm.

  16. Orthogonal polynomial approximation in higher dimensions: Applications in astrodynamics

    NASA Astrophysics Data System (ADS)

    Bani Younes, Ahmad Hani Abd Alqader

    We propose novel methods to utilize orthogonal polynomial approximation in higher dimension spaces, which enable us to modify classical differential equation solvers to perform high precision, long-term orbit propagation. These methods have immediate application to efficient propagation of catalogs of Resident Space Objects (RSOs) and improved accounting for the uncertainty in the ephemeris of these objects. More fundamentally, the methodology promises to be of broad utility in solving initial and two point boundary value problems from a wide class of mathematical representations of problems arising in engineering, optimal control, physical sciences and applied mathematics. We unify and extend classical results from function approximation theory and consider their utility in astrodynamics. Least square approximation, using the classical Chebyshev polynomials as basis functions, is reviewed for discrete samples of the to-be-approximated function. We extend the orthogonal approximation ideas to n-dimensions in a novel way, through the use of array algebra and Kronecker operations. Approximation of test functions illustrates the resulting algorithms and provides insight into the errors of approximation, as well as the associated errors arising when the approximations are differentiated or integrated. Two sets of applications are considered that are challenges in astrodynamics. The first application addresses local approximation of high degree and order geopotential models, replacing the global spherical harmonic series by a family of locally precise orthogonal polynomial approximations for efficient computation. A method is introduced which adapts the approximation degree radially, compatible with the truth that the highest degree approximations (to ensure maximum acceleration error < 10-9 ms-2, globally) are required near the Earths surface, whereas lower degree approximations are required as radius increases. We show that a four order of magnitude speedup is

  17. Transfer Function Identification Using Orthogonal Fourier Transform Modeling Functions

    NASA Technical Reports Server (NTRS)

    Morelli, Eugene A.

    2013-01-01

    A method for transfer function identification, including both model structure determination and parameter estimation, was developed and demonstrated. The approach uses orthogonal modeling functions generated from frequency domain data obtained by Fourier transformation of time series data. The method was applied to simulation data to identify continuous-time transfer function models and unsteady aerodynamic models. Model fit error, estimated model parameters, and the associated uncertainties were used to show the effectiveness of the method for identifying accurate transfer function models from noisy data.

  18. Bacillus subtilis GlnR contains an autoinhibitory C-terminal domain required for the interaction with glutamine synthetase.

    PubMed

    Wray, Lewis V; Fisher, Susan H

    2008-04-01

    The Bacillus subtilis GlnR transcription factor regulates gene expression in response to changes in nitrogen availability. Glutamine synthetase transmits the nitrogen regulatory signal to GlnR. The DNA-binding activity of GlnR is activated by a transient protein-protein interaction with feedback-inhibited glutamine synthetase that stabilizes GlnR-DNA complexes. This signal transduction mechanism was analysed by creating mutant GlnR proteins with partial or complete truncations of their C-terminal domains. The truncated GlnR proteins were found to constitutively repress gene expression in vivo. This constitutive repression did not require glutamine synthetase. Purified mutant GlnR proteins bound DNA in vitro more tightly than wild-type GlnR protein and this binding was not activated by feedback-inhibited glutamine synthetase. While full-length GlnR is monomeric, the truncated GlnR proteins contained significant levels of dimers. These results indicate that the C-terminal region of GlnR acts as an autoinhibitory domain that prevents GlnR dimerization and thus impedes DNA binding. The GlnR C-terminal domain is also required for the interaction between GlnR and feedback-inhibited glutamine synthetase. Compared with the full-length GlnR protein, the truncated GlnR proteins were defective in their interaction with feedback-inhibited glutamine synthetase in cross-linking experiments.

  19. A CMP-N-acetylneuraminic acid synthetase purified from a marine bacterium, Photobacterium leiognathi JT-SHIZ-145.

    PubMed

    Kajiwara, Hitomi; Mine, Toshiki; Miyazaki, Tatsuo; Yamamoto, Takeshi

    2011-01-01

    A cytidine 5'-monophospho-N-acetylneuraminic acid (CMP-Neu5Ac) synthetase was found in a crude extract prepared from Photobacterium leiognathi JT-SHIZ-145, a marine bacterium that also produces a β-galactoside α2,6-sialyltransferase. The CMP-Neu5Ac synthetase was purified from the crude extract of the cells by a combination of anion-exchange and gel filtration column chromatography. The purified enzyme migrated as a single band (60 kDa) on sodium dodecylsulfate-polyacrylamide gel electrophoresis. The activity of the enzyme was maximal at 35 °C at pH 9.0, and the synthetase required Mg(2+) for activity. Although these properties are similar to those of other CMP-Neu5Ac synthetases isolated from bacteria, this synthetase produced not only CMP-Neu5Ac from cytidine triphosphate and Neu5Ac, but also CMP-N-glycolylneuraminic acid from cytidine triphosphate and N-glycolylneuraminic acid, unlike CMP-Neu5Ac synthetase purified from Escherichia coli.

  20. The purification and properties of the glutamine synthetase from the cytosol of Soya-bean root nodules.

    PubMed Central

    McParland, R H; Guevara, J G; Becker, R R; Evans, H J

    1976-01-01

    The major portion of glutamine synthetase activity in root nodules of soya-bean plants is associated with the cytosol rather than with Rhizobium japonicum bacteroids. Glutamine synthetase accounts for about 2% of the total soluble protein in nodule cytosol. Glutamine synthetase from nodule cytosol has been purified by a procedure involving fractionation with protamine sulphate, ammonium sulphate and polypropylene glycol, chromatography on DEAE-Bio-Gel A and Bio-Gel A-5m and affinity chromatography on glutamate-agarose columns. The purified preparation appeared to be homogeneous in the analytical ultracentrifuge. From sedimentation-equilibrium experiments a mol. wt. of about 376000 was determined for the native enzyme and 47300 for the enzyme in guanidinium chloride. From these data and measurements of electron micrographs, we have concluded that glutamine synthetase from nodule cytosol consists of eight subunits arranged in two sets of planar tetramers which form a cubical configuration with dimensions of about 10 nm (100 A) across each side. Glutamine synthetase from nodule cytosol has a higher glycine and proline content and a lower content of phenylalanine than the glutamine synthetase that has been prepared from pea seed. The cytosol enzyme contains four half-cystine molecules per subunit, which is in contrast with two reported for the enzyme from pea seed. Enzyme activity is striking influenced by the relative proportion of Mg2+ and Mn2+ in the assay medium. Activity is inhibited by feedback inhibitors and is influenced by energy charge. Images PLATE 1 PLATE 2 PMID:8035

  1. Proofreading in vivo: Editing of homocysteine by methionyl-tRNA synthetase in Escherichia coli

    SciTech Connect

    Jakubowski, H. )

    1990-06-01

    Previous in vitro studies have established a pre-transfer proofreading mechanism for editing of homocysteine by bacterial methionyl-, isoleucyl-, and valyl-tRNA synthetases. The unusual feature of the editing is the formation of a distinct compound, homocysteine thiolactone. Now, two-dimensional TLC analysis of 35S-labeled amino acids extracted from cultures of the bacterium Escherichia coli reveals that the thiolactone is also synthesized in vivo. In E. coli, the thiolactone is made from homocysteine in a reaction catalyzed by methionyl-tRNA synthetase. One molecule of homocysteine is edited as thiolactone per 109 molecules of methionine incorporated into protein in vivo. These results not only directly demonstrate that the adenylate proofreading pathway for rejection of misactivated homocysteine operates in vivo in E. coli but, in general, establish the importance of error-editing mechanisms in living cells.

  2. Aminoacyl-tRNA synthetase dependent angiogenesis revealed by a bioengineered macrolide inhibitor.

    PubMed

    Mirando, Adam C; Fang, Pengfei; Williams, Tamara F; Baldor, Linda C; Howe, Alan K; Ebert, Alicia M; Wilkinson, Barrie; Lounsbury, Karen M; Guo, Min; Francklyn, Christopher S

    2015-08-14

    Aminoacyl-tRNA synthetases (AARSs) catalyze an early step in protein synthesis, but also regulate diverse physiological processes in animal cells. These include angiogenesis, and human threonyl-tRNA synthetase (TARS) represents a potent pro-angiogenic AARS. Angiogenesis stimulation can be blocked by the macrolide antibiotic borrelidin (BN), which exhibits a broad spectrum toxicity that has discouraged deeper investigation. Recently, a less toxic variant (BC194) was identified that potently inhibits angiogenesis. Employing biochemical, cell biological, and biophysical approaches, we demonstrate that the toxicity of BN and its derivatives is linked to its competition with the threonine substrate at the molecular level, which stimulates amino acid starvation and apoptosis. By separating toxicity from the inhibition of angiogenesis, a direct role for TARS in vascular development in the zebrafish could be demonstrated. Bioengineered natural products are thus useful tools in unmasking the cryptic functions of conventional enzymes in the regulation of complex processes in higher metazoans.

  3. Aminoacyl transfer ribonucleic acid synthetases from cell-free extract of Plasmodium berghei.

    PubMed

    Ilan, J; Ilan, J

    1969-05-02

    Aminoacyl transfer ribonucleic acid synthetases for leucine tyrosine, histidine, valine, proline, threonine, and lysine were obtainnned from cell-free extract of Plasmodium berghei. The leucyl-tRNA synthetase cane charge tRNA from liver or Escherichia coli with leucine-c(14), liver tRNA being a better substrate. The amount of aminoacylation increses linerly with respect to the quantity of tRNA added from either source and is dependent on the amount of enzyme added. The rate of aminoacylation is constant for 10 minutes and then decreases. It is enhanced by polyvinylsulfate. One-tenth millimoler pyrimethamine, hydroxystilbamidine, quinacrine, and acriflavine inhibited the formation of C(14)-valyl-tRNA. Species specificity between tRNA and its charging enzyme with respect to the recognition site is discussed.

  4. Human lysyl-tRNA synthetase is secreted to trigger proinflammatory response

    PubMed Central

    Park, Sang Gyu; Kim, Hye Jin; Min, You Hong; Choi, Eung-Chil; Shin, Young Kee; Park, Bum-Joon; Lee, Sang Won; Kim, Sunghoon

    2005-01-01

    Although aminoacyl-tRNA synthetases (ARSs) are essential for protein synthesis, they also function as regulators and signaling molecules in diverse biological processes. Here, we screened 11 different human ARSs to identify the enzyme that is secreted as a signaling molecule. Among them, we found that lysyl-tRNA synthetase (KRS) was secreted from intact human cells, and its secretion was induced by TNF-α. The secreted KRS bound to macrophages and peripheral blood mononuclear cells to enhance the TNF-α production and their migration. The mitogen-activated protein kinases, extracellular signal-regulated kinase and p38 mitogen-activated protein kinase, and Gαi were determined to be involved in the signal transduction triggered by KRS. All of these activities demonstrate that human KRS may work as a previously uncharacterized signaling molecule, inducing immune response through the activation of monocyte/macrophages. PMID:15851690

  5. let-65 is cytoplasmic methionyl tRNA synthetase in C. elegans

    PubMed Central

    Alriyami, Maha Z.; Jones, Martin R.; Johnsen, Robert C.; Banerjee, Yajnavalka; Baillie, David L.

    2014-01-01

    Cytoplasmic methionyl tRNA synthetase (MetRS) is one of more than 20 cytoplasmic aminoacyl tRNA synthetase enzymes (ARS). This family of enzymes catalyzes a process fundamental for protein translation. Using a combination of genetic mapping, oligonucleotide array comparative genomic hybridization, and phenotypic correlation, we show that mutations in the essential gene, let-65, reside within the predicted Caenorhabditis elegans homologue of MetRS, which we have named mars-1. We demonstrate that the lethality associated with alleles of let-65 is fully rescued by a transgenic array that spans the mars-1 genomic region. Furthermore, sequence analysis reveals that six let-65 alleles lead to the alteration of highly conserved amino acids. PMID:25606464

  6. Genetic incorporation of histidine derivatives using an engineered pyrrolysyl-tRNA synthetase.

    PubMed

    Xiao, Han; Peters, Francis B; Yang, Peng-Yu; Reed, Sean; Chittuluru, Johnathan R; Schultz, Peter G

    2014-05-16

    A polyspecific amber suppressor aminoacyl-tRNA synthetase/tRNA pair was evolved that genetically encodes a series of histidine analogues in both Escherichia coli and mammalian cells. In combination with tRNACUA(Pyl), a pyrrolysyl-tRNA synthetase mutant was able to site-specifically incorporate 3-methyl-histidine, 3-pyridyl-alanine, 2-furyl-alanine, and 3-(2-thienyl)-alanine into proteins in response to an amber codon. Substitution of His66 in the blue fluorescent protein (BFP) with these histidine analogues created mutant proteins with distinct spectral properties. This work further expands the structural and chemical diversity of unnatural amino acids (UAAs) that can be genetically encoded in prokaryotic and eukaryotic organisms and affords new probes of protein structure and function.

  7. Proofreading in vivo: editing of homocysteine by methionyl-tRNA synthetase in Escherichia coli.

    PubMed Central

    Jakubowski, H

    1990-01-01

    Previous in vitro studies have established a pre-transfer proofreading mechanism for editing of homocysteine by bacterial methionyl-, isoleucyl-, and valyl-tRNA synthetases. The unusual feature of the editing is the formation of a distinct compound, homocysteine thiolactone. Now, two-dimensional TLC analysis of 35S-labeled amino acids extracted from cultures of the bacterium Escherichia coli reveals that the thiolactone is also synthesized in vivo. In E. coli, the thiolactone is made from homocysteine in a reaction catalyzed by methionyl-tRNA synthetase. One molecule of homocysteine is edited as thiolactone per 109 molecules of methionine incorporated into protein in vivo. These results not only directly demonstrate that the adenylate proofreading pathway for rejection of misactivated homocysteine operates in vivo in E. coli but, in general, establish the importance of error-editing mechanisms in living cells. Images PMID:2191291

  8. Multistep modeling of protein structure: application towards refinement of tyr-tRNA synthetase

    NASA Technical Reports Server (NTRS)

    Srinivasan, S.; Shibata, M.; Roychoudhury, M.; Rein, R.

    1987-01-01

    The scope of multistep modeling (MSM) is expanding by adding a least-squares minimization step in the procedure to fit backbone reconstruction consistent with a set of C-alpha coordinates. The analytical solution of Phi and Psi angles, that fits a C-alpha x-ray coordinate is used for tyr-tRNA synthetase. Phi and Psi angles for the region where the above mentioned method fails, are obtained by minimizing the difference in C-alpha distances between the computed model and the crystal structure in a least-squares sense. We present a stepwise application of this part of MSM to the determination of the complete backbone geometry of the 321 N terminal residues of tyrosine tRNA synthetase to a root mean square deviation of 0.47 angstroms from the crystallographic C-alpha coordinates.

  9. Effects of aeration on formation and localization of the acetyl coenzyme A synthetases of Saccharomyces cerevisiae

    NASA Technical Reports Server (NTRS)

    Klein, H. P.; Jahnke, L.

    1979-01-01

    Previous studies on the yeast Saccharomyces cerevisiae have shown that two different forms of the enzyme acetyl coenzyme A synthetase (ACS) are present, depending on the conditions under which the cells are grown. The paper evaluates the usefulness of a method designed to assay both synthetases simultaneously in yeast homogenates. The data presented confirm the possibility of simultaneous detection and estimation of the amount of both ACSs of S. cerevisiae in crude homogenates of this strain, making possible the study of physiological factors involved in the formation of these isoenzymes. One important factor for specifying which of the two enzymes is found in these yeast cells is the presence or absence of oxygen in their environment. Aeration not only affects the ratio of the two ACSs but also appears to affect the cellular distribution of these enzymes. Most of the data presented suggest the possibility that the nonaerobic ACS may serve as a precursor to the aerobic form.

  10. Assessing the effects of threonyl-tRNA synthetase on angiogenesis-related responses.

    PubMed

    Mirando, Adam C; Abdi, Khadar; Wo, Peibin; Lounsbury, Karen M

    2017-01-15

    Several recent reports have found a connection between specific aminoacyl-tRNA synthetases and the regulation of angiogenesis. As this new area of research is explored, it is important to have reliable assays to assess the specific angiogenesis functions of these enzymes. This review provides information about specific in vitro and in vivo methods that were used to assess the angiogenic functions of threonyl-tRNA synthetase including endothelial cell migration and tube assays as well as chorioallantoic membrane and tumor vascularization assays. The theory and discussion include best methods of analysis and quantification along with the advantages and limitations of each type of assay. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Effects of aeration on formation and localization of the acetyl coenzyme A synthetases of Saccharomyces cerevisiae

    NASA Technical Reports Server (NTRS)

    Klein, H. P.; Jahnke, L.

    1979-01-01

    Previous studies on the yeast Saccharomyces cerevisiae have shown that two different forms of the enzyme acetyl coenzyme A synthetase (ACS) are present, depending on the conditions under which the cells are grown. The paper evaluates the usefulness of a method designed to assay both synthetases simultaneously in yeast homogenates. The data presented confirm the possibility of simultaneous detection and estimation of the amount of both ACSs of S. cerevisiae in crude homogenates of this strain, making possible the study of physiological factors involved in the formation of these isoenzymes. One important factor for specifying which of the two enzymes is found in these yeast cells is the presence or absence of oxygen in their environment. Aeration not only affects the ratio of the two ACSs but also appears to affect the cellular distribution of these enzymes. Most of the data presented suggest the possibility that the nonaerobic ACS may serve as a precursor to the aerobic form.

  12. Aminoacyl-tRNA synthetases: versatile players in the changing theater of translation.

    PubMed Central

    Francklyn, Christopher; Perona, John J; Puetz, Joern; Hou, Ya-Ming

    2002-01-01

    Aminoacyl-tRNA synthetases attach amino acids to the 3' termini of cognate tRNAs to establish the specificity of protein synthesis. A recent Asilomar conference (California, January 13-18, 2002) discussed new research into the structure-function relationship of these crucial enzymes, as well as a multitude of novel functions, including participation in amino acid biosynthesis, cell cycle control, RNA splicing, and export of tRNAs from nucleus to cytoplasm in eukaryotic cells. Together with the discovery of their role in the cellular synthesis of proteins to incorporate selenocysteine and pyrrolysine, these diverse functions of aminoacyl-tRNA synthetases underscore the flexibility and adaptability of these ancient enzymes and stimulate the development of new concepts and methods for expanding the genetic code. PMID:12458790

  13. Structure of Human Phosphopantothenoylcysteine Synthetase at 2.3 Å Resolution

    SciTech Connect

    Manoj, N.; Strauss, E.; Begley, T.P.; Ealick, S.E.

    2010-12-01

    The structure of human phosphopantothenoylcysteine (PPC) synthetase was determined at 2.3 {angstrom} resolution. PPC synthetase is a dimer with identical monomers. Some features of the monomer fold resemble a group of NAD-dependent enzymes, while other features resemble the ribokinase fold. The ATP, phosphopantothenate, and cysteine binding sites were deduced from modeling studies. Highly conserved ATP binding residues include Gly43, Ser61, Gly63, Gly66, Phe230, and Asn258. Highly conserved phosphopantothenate binding residues include Asn59, Ala179, Ala180, and Asp183 from one monomer and Arg55 from the adjacent monomer. The structure predicts a ping pong mechanism with initial formation of an acyladenylate intermediate, followed by release of pyrophosphate and attack by cysteine to form the final products PPC and AMP.

  14. Multistep modeling of protein structure: application towards refinement of tyr-tRNA synthetase

    NASA Technical Reports Server (NTRS)

    Srinivasan, S.; Shibata, M.; Roychoudhury, M.; Rein, R.

    1987-01-01

    The scope of multistep modeling (MSM) is expanding by adding a least-squares minimization step in the procedure to fit backbone reconstruction consistent with a set of C-alpha coordinates. The analytical solution of Phi and Psi angles, that fits a C-alpha x-ray coordinate is used for tyr-tRNA synthetase. Phi and Psi angles for the region where the above mentioned method fails, are obtained by minimizing the difference in C-alpha distances between the computed model and the crystal structure in a least-squares sense. We present a stepwise application of this part of MSM to the determination of the complete backbone geometry of the 321 N terminal residues of tyrosine tRNA synthetase to a root mean square deviation of 0.47 angstroms from the crystallographic C-alpha coordinates.

  15. Aminoacyl-tRNA synthetase dependent angiogenesis revealed by a bioengineered macrolide inhibitor

    PubMed Central

    Mirando, Adam C.; Fang, Pengfei; Williams, Tamara F.; Baldor, Linda C.; Howe, Alan K.; Ebert, Alicia M.; Wilkinson, Barrie; Lounsbury, Karen M.; Guo, Min; Francklyn, Christopher S.

    2015-01-01

    Aminoacyl-tRNA synthetases (AARSs) catalyze an early step in protein synthesis, but also regulate diverse physiological processes in animal cells. These include angiogenesis, and human threonyl-tRNA synthetase (TARS) represents a potent pro-angiogenic AARS. Angiogenesis stimulation can be blocked by the macrolide antibiotic borrelidin (BN), which exhibits a broad spectrum toxicity that has discouraged deeper investigation. Recently, a less toxic variant (BC194) was identified that potently inhibits angiogenesis. Employing biochemical, cell biological, and biophysical approaches, we demonstrate that the toxicity of BN and its derivatives is linked to its competition with the threonine substrate at the molecular level, which stimulates amino acid starvation and apoptosis. By separating toxicity from the inhibition of angiogenesis, a direct role for TARS in vascular development in the zebrafish could be demonstrated. Bioengineered natural products are thus useful tools in unmasking the cryptic functions of conventional enzymes in the regulation of complex processes in higher metazoans. PMID:26271225

  16. Purification, crystallization and data collection of methicillin-resistant Staphylococcus aureus Sar2676, a pantothenate synthetase

    PubMed Central

    Seetharamappa, Jaldappagari; Oke, Muse; Liu, Huanting; McMahon, Stephen A.; Johnson, Kenneth A.; Carter, Lester; Dorward, Mark; Zawadzki, Michal; Overton, Ian M.; van Niekirk, C. A. Johannes; Graham, Shirley; Botting, Catherine H.; Taylor, Garry L.; White, Malcolm F.; Barton, Geoffrey J.; Coote, Peter J.; Naismith, James H.

    2007-01-01

    Sar2676, a pantothenate synthetase with a molecular weight of 31 419 Da from methicillin-resistant Staphylococcus aureus, has been expressed, purified and crystallized at 293 K. The protein crystallizes in a primitive triclinic lattice, with unit-cell parameters a = 45.3, b = 60.5, c = 117.6 Å, α = 87.2, β = 81.2, γ = 68.4°. A complete data set has been collected to 2.3 Å resolution at the ESRF. Consideration of the likely solvent content suggested the asymmetric unit to contain four molecules. This has been confirmed by molecular-replacement phasing calculations, which give a solution with four monomers using a monomer of pantothenate synthetase from Escherichia coli (PDB code 1iho), which is 41% identical to Sar2676, as a search model. PMID:17554169

  17. Reassimilation of Photorespiratory Ammonium in Lotus japonicus Plants Deficient in Plastidic Glutamine Synthetase.

    PubMed

    Pérez-Delgado, Carmen M; García-Calderón, Margarita; Márquez, Antonio J; Betti, Marco

    2015-01-01

    It is well established that the plastidic isoform of glutamine synthetase (GS2) is the enzyme in charge of photorespiratory ammonium reassimilation in plants. The metabolic events associated to photorespiratory NH4(+) accumulation were analyzed in a Lotus japonicus photorespiratory mutant lacking GS2. The mutant plants accumulated high levels of NH4(+) when photorespiration was active, followed by a sudden drop in the levels of this compound. In this paper it was examined the possible existence of enzymatic pathways alternative to GS2 that could account for this decline in the photorespiratory ammonium. Induction of genes encoding for cytosolic glutamine synthetase (GS1), glutamate dehydrogenase (GDH) and asparagine synthetase (ASN) was observed in the mutant in correspondence with the diminishment of NH4(+). Measurements of gene expression, polypeptide levels, enzyme activity and metabolite levels were carried out in leaf samples from WT and mutant plants after different periods of time under active photorespiratory conditions. In the case of asparagine synthetase it was not possible to determine enzyme activity and polypeptide content; however, an increased asparagine content in parallel with the induction of ASN gene expression was detected in the mutant plants. This increase in asparagine levels took place concomitantly with an increase in glutamine due to the induction of cytosolic GS1 in the mutant, thus revealing a major role of cytosolic GS1 in the reassimilation and detoxification of photorespiratory NH4(+) when the plastidic GS2 isoform is lacking. Moreover, a diminishment in glutamate levels was observed, that may be explained by the induction of NAD(H)-dependent GDH activity.

  18. Thiol synthetases of legumes: immunogold localization and differential gene regulation by phytohormones.

    PubMed

    Clemente, Maria R; Bustos-Sanmamed, Pilar; Loscos, Jorge; James, Euan K; Pérez-Rontomé, Carmen; Navascués, Joaquín; Gay, Marina; Becana, Manuel

    2012-06-01

    In plants and other organisms, glutathione (GSH) biosynthesis is catalysed sequentially by γ-glutamylcysteine synthetase (γECS) and glutathione synthetase (GSHS). In legumes, homoglutathione (hGSH) can replace GSH and is synthesized by γECS and a specific homoglutathione synthetase (hGSHS). The subcellular localization of the enzymes was examined by electron microscopy in several legumes and gene expression was analysed in Lotus japonicus plants treated for 1-48 h with 50 μM of hormones. Immunogold localization studies revealed that γECS is confined to chloroplasts and plastids, whereas hGSHS is also in the cytosol. Addition of hormones caused differential expression of thiol synthetases in roots. After 24-48 h, abscisic and salicylic acids downregulated GSHS whereas jasmonic acid upregulated it. Cytokinins and polyamines activated GSHS but not γECS or hGSHS. Jasmonic acid elicited a coordinated response of the three genes and auxin induced both hGSHS expression and activity. Results show that the thiol biosynthetic pathway is compartmentalized in legumes. Moreover, the similar response profiles of the GSH and hGSH contents in roots of non-nodulated and nodulated plants to the various hormonal treatments indicate that thiol homeostasis is independent of the nitrogen source of the plants. The differential regulation of the three mRNA levels, hGSHS activity, and thiol contents by hormones indicates a fine control of thiol biosynthesis at multiple levels and strongly suggests that GSH and hGSH play distinct roles in plant development and stress responses.

  19. Thiol synthetases of legumes: immunogold localization and differential gene regulation by phytohormones

    PubMed Central

    Clemente, Maria R.; Bustos-Sanmamed, Pilar; Loscos, Jorge; James, Euan K.; Pérez-Rontomé, Carmen; Navascués, Joaquín; Gay, Marina; Becana, Manuel

    2012-01-01

    In plants and other organisms, glutathione (GSH) biosynthesis is catalysed sequentially by γ-glutamylcysteine synthetase (γECS) and glutathione synthetase (GSHS). In legumes, homoglutathione (hGSH) can replace GSH and is synthesized by γECS and a specific homoglutathione synthetase (hGSHS). The subcellular localization of the enzymes was examined by electron microscopy in several legumes and gene expression was analysed in Lotus japonicus plants treated for 1–48 h with 50 μM of hormones. Immunogold localization studies revealed that γECS is confined to chloroplasts and plastids, whereas hGSHS is also in the cytosol. Addition of hormones caused differential expression of thiol synthetases in roots. After 24–48 h, abscisic and salicylic acids downregulated GSHS whereas jasmonic acid upregulated it. Cytokinins and polyamines activated GSHS but not γECS or hGSHS. Jasmonic acid elicited a coordinated response of the three genes and auxin induced both hGSHS expression and activity. Results show that the thiol biosynthetic pathway is compartmentalized in legumes. Moreover, the similar response profiles of the GSH and hGSH contents in roots of non-nodulated and nodulated plants to the various hormonal treatments indicate that thiol homeostasis is independent of the nitrogen source of the plants. The differential regulation of the three mRNA levels, hGSHS activity, and thiol contents by hormones indicates a fine control of thiol biosynthesis at multiple levels and strongly suggests that GSH and hGSH play distinct roles in plant development and stress responses. PMID:22442424

  20. Quality Control by Isoleucyl-tRNA Synthetase of Bacillus subtilis Is Required for Efficient Sporulation

    PubMed Central

    Kermgard, Elizabeth; Yang, Zhou; Michel, Annika-Marisa; Simari, Rachel; Wong, Jacqueline; Ibba, Michael; Lazazzera, Beth A.

    2017-01-01

    Isoleucyl-tRNA synthetase (IleRS) is an aminoacyl-tRNA synthetase whose essential function is to aminoacylate tRNAIle with isoleucine. Like some other aminoacyl-tRNA synthetases, IleRS can mischarge tRNAIle and correct this misacylation through a separate post-transfer editing function. To explore the biological significance of this editing function, we created a ileS(T233P) mutant of Bacillus subtilis that allows tRNAIle mischarging while retaining wild-type Ile-tRNAIle synthesis activity. As seen in other species defective for aminoacylation quality control, the growth rate of the ileS(T233P) strain was not significantly different from wild-type. When the ileS(T233P) strain was assessed for its ability to promote distinct phenotypes in response to starvation, the ileS(T233P) strain was observed to exhibit a significant defect in formation of environmentally resistant spores. The sporulation defect ranged from 3-fold to 30-fold and was due to a delay in activation of early sporulation genes. The loss of aminoacylation quality control in the ileS(T233P) strain resulted in the inability to compete with a wild-type strain under selective conditions that required sporulation. These data show that the quality control function of IleRS is required in B. subtilis for efficient sporulation and suggests that editing by aminoacyl-tRNA synthetases may be important for survival under starvation/nutrient limitation conditions. PMID:28139725

  1. Glutamine synthetase immunor present in oligodendroglia of regions of the central nervous system

    NASA Technical Reports Server (NTRS)

    D'Amelio, Fernando; Eng, Lawrence F.; Gibbs, Michael A.

    1990-01-01

    Glutamine synthetase immunoreactive oligodendrocytes were identified in the cerebral cortex, cerebellum, brain stem, and spinal cord. They were mostly confined to the gray matter, particularly close to neurons and processes. The white matter showed few immunoreactive oligodendroglia. It was suggested that some type of oligodendrocytes, specially those in perineuronal location, might fulfill a functional role more akin to astrocytes than to the normally myelinating oligodendroglia.

  2. A Phenotypic Based Target Screening Approach Delivers New Antitubercular CTP Synthetase Inhibitors.

    PubMed

    Esposito, Marta; Szadocka, Sára; Degiacomi, Giulia; Orena, Beatrice S; Mori, Giorgia; Piano, Valentina; Boldrin, Francesca; Zemanová, Júlia; Huszár, Stanislav; Barros, David; Ekins, Sean; Lelièvre, Joel; Manganelli, Riccardo; Mattevi, Andrea; Pasca, Maria Rosalia; Riccardi, Giovanna; Ballell, Lluis; Mikušová, Katarína; Chiarelli, Laurent R

    2017-06-09

    Despite its great potential, the target-based approach has been mostly unsuccessful in tuberculosis drug discovery, while whole cell phenotypic screening has delivered several active compounds. However, for many of these hits, the cellular target has not yet been identified, thus preventing further target-based optimization of the compounds. In this context, the newly validated drug target CTP synthetase PyrG was exploited to assess a target-based approach of already known, but untargeted, antimycobacterial compounds. To this purpose the publically available GlaxoSmithKline antimycobacterial compound set was assayed, uncovering a series of 4-(pyridin-2-yl)thiazole derivatives which efficiently inhibit the Mycobacterium tuberculosis PyrG enzyme activity, one of them showing low activity against the human CTP synthetase. The three best compounds were ATP binding site competitive inhibitors, with Ki values ranging from 3 to 20 μM, but did not show any activity against a small panel of different prokaryotic and eukaryotic kinases, thus demonstrating specificity for the CTP synthetases. Metabolic labeling experiments demonstrated that the compounds directly interfere not only with CTP biosynthesis, but also with other CTP dependent biochemical pathways, such as lipid biosynthesis. Moreover, using a M. tuberculosis pyrG conditional knock-down strain, it was shown that the activity of two compounds is dependent on the intracellular concentration of the CTP synthetase. All these results strongly suggest a role of PyrG as a target of these compounds, thus strengthening the value of this kind of approach for the identification of new scaffolds for drug development.

  3. Glutamine synthetase immunoreactivity is present in oligodendroglia of various regions of the central nervous system

    NASA Technical Reports Server (NTRS)

    D'Amelio, F.; Eng, L. F.; Gibbs, M. A.

    1990-01-01

    Glutamine synthetase immunoreactive oligodendrocytes were identified in the cerebral cortex, cerebellum, brain stem, and spinal cord. They were mostly confined to the gray matter, particularly close to neurons and processes. The white matter showed few immunoreactive oligodendroglia. It was suggested that some type of oligodendrocytes, specially those in perineuronal location, might fulfill a functional role more akin to astrocytes than to the normally myelinating oligodendroglia.

  4. Co-operation between Polymerases and Nucleotide Synthetases in the RNA World

    PubMed Central

    Kim, Ye Eun; Higgs, Paul G.

    2016-01-01

    It is believed that life passed through an RNA World stage in which replication was sustained by catalytic RNAs (ribozymes). The two most obvious types of ribozymes are a polymerase, which uses a neighbouring strand as a template to make a complementary sequence to the template, and a nucleotide synthetase, which synthesizes monomers for use by the polymerase. When a chemical source of monomers is available, the polymerase can survive on its own. When the chemical supply of monomers is too low, nucleotide production by the synthetase is essential and the two ribozymes can only survive when they are together. Here we consider a computational model to investigate conditions under which coexistence and cooperation of these two types of ribozymes is possible. The model considers six types of strands: the two functional sequences, the complementary strands to these sequences (which are required as templates), and non-functional mutants of the two sequences (which act as parasites). Strands are distributed on a two-dimensional lattice. Polymerases replicate strands on neighbouring sites and synthetases produce monomers that diffuse in the local neighbourhood. We show that coexistence of unlinked polymerases and synthetases is possible in this spatial model under conditions in which neither sequence could survive alone; hence, there is a selective force for increasing complexity. Coexistence is dependent on the relative lengths of the two functional strands, the strand diffusion rate, the monomer diffusion rate, and the rate of deleterious mutations. The sensitivity of this two-ribozyme system suggests that evolution of a system of many types of ribozymes would be difficult in a purely spatial model with unlinked genes. We therefore speculate that linkage of genes onto mini-chromosomes and encapsulation of strands in protocells would have been important fairly early in the history of life as a means of enabling more complex systems to evolve. PMID:27820829

  5. What is the oligoadenylate synthetases-like protein and does it have therapeutic potential for influenza?

    PubMed

    Alcorn, John F; Sarkar, Saumendra N

    2015-02-01

    Besides its pandemic potential, seasonal influenza infection is associated with an estimated 250,000 to 500,000 deaths worldwide every year. Part of this virulence of influenza virus can be attributed to its ability to evade the host innate immune response. Here, we discuss the possibility of using a recently described mechanism of boosting the innate immunity by oligoadenylate synthetase-like protein, to combat influenza infections.

  6. Glutamine synthetase gene expression and glutamate transporters in C6-glioma cells.

    PubMed

    Baber, Zafeer; Haghighat, Nasrin

    2010-12-01

    Glutamine synthetase (GS) is the major glutamate-forming enzyme of vertebrae and is accepted to be a marker of astroglial cells. Maturation of astroglial cells is characterized by an increase in GS activity, and the regulation of this enzyme is the topic of many publications. The amino acid glutamate is the major excitatory neurotransmitter in the brain and mediates normal excitatory synaptic transmission by interaction with postsynaptic receptors. Glutamate also acts as a potent neurotoxin when present at high concentration. Glutamate neurotoxicity plays an important role in the pathophysiology of many neurological disorders, such as Alzheimer's disease, Huntington's disease and amyotrophic lateral sclerosis. In the normal condition, L-glutamate is predominantly taken up, metabolized and recycled by astrocytes through the glutamate transporters (GLAST/GLT1) and glutamine synthetase (GS) catalytic activity. Because of the fundamental role of these glutamate transporters and the glutamine synthetase enzyme in controlling cerebral glutamate level, regulation of GS and studying of the glutamate transporters in glial cells is important. Astrocytes are supportive cells and act as the site of detoxification of glutamate in the brain. However, their isolation from the brain is a tedious, costly and time consuming procedure. On the other hand, the C6-glioma cells are readily available on the market. They are well characterized and have been a useful model for CNS glia in many laboratories. For this study, we used the C6-glioma cell line as a model system. We examined the presence or absence of glial specific glutamate transporters (GLTI and GLAST) in C6-glioma cells, which was done by immunocytochemistry. We also examined glutamine synthetase gene expression in these cells by treatment of the C6-glioma cells with estrogen (17ß estradiol). The findings from this study provide useful information about C6-glioma cells which makes the study of the CNS tremendously inexpensive.

  7. Mitochondrial phenylalanyl-tRNA synthetase mutations underlie fatal infantile Alpers encephalopathy.

    PubMed

    Elo, Jenni M; Yadavalli, Srujana S; Euro, Liliya; Isohanni, Pirjo; Götz, Alexandra; Carroll, Christopher J; Valanne, Leena; Alkuraya, Fowzan S; Uusimaa, Johanna; Paetau, Anders; Caruso, Eric M; Pihko, Helena; Ibba, Michael; Tyynismaa, Henna; Suomalainen, Anu

    2012-10-15

    Next-generation sequencing has turned out to be a powerful tool to uncover genetic basis of childhood mitochondrial disorders. We utilized whole-exome analysis and discovered novel compound heterozygous mutations in FARS2 (mitochondrial phenylalanyl transfer RNA synthetase), encoding the mitochondrial phenylalanyl transfer RNA (tRNA) synthetase (mtPheRS) in two patients with fatal epileptic mitochondrial encephalopathy. The mutations affected highly conserved amino acids, p.I329T and p.D391V. Recently, a homozygous FARS2 variant p.Y144C was reported in a Saudi girl with mitochondrial encephalopathy, but the pathogenic role of the variant remained open. Clinical features, including postnatal onset, catastrophic epilepsy, lactic acidemia, early lethality and neuroimaging findings of the patients with FARS2 variants, resembled each other closely, and neuropathology was consistent with Alpers syndrome. Our structural analysis of mtPheRS predicted that p.I329T weakened ATP binding in the aminoacylation domain, and in vitro studies with recombinant mutant protein showed decreased affinity of this variant to ATP. Furthermore, p.D391V and p.Y144C were predicted to disrupt synthetase function by interrupting the rotation of the tRNA anticodon stem-binding domain from a closed to an open form. In vitro characterization indicated reduced affinity of p.D391V mutant protein to phenylalanine, whereas p.Y144C disrupted tRNA binding. The stability of p.I329T and p.D391V mutants in a refolding assay was impaired. Our results imply that the three FARS2 mutations directly impair aminoacylation function and stability of mtPheRS, leading to a decrease in overall tRNA charging capacity. This study establishes a new genetic cause of infantile mitochondrial Alpers encephalopathy and reports a new mitochondrial aminoacyl-tRNA synthetase as a cause of mitochondrial disease.

  8. Glutamine synthetase immunoreactivity is present in oligodendroglia of various regions of the central nervous system

    NASA Technical Reports Server (NTRS)

    D'Amelio, F.; Eng, L. F.; Gibbs, M. A.

    1990-01-01

    Glutamine synthetase immunoreactive oligodendrocytes were identified in the cerebral cortex, cerebellum, brain stem, and spinal cord. They were mostly confined to the gray matter, particularly close to neurons and processes. The white matter showed few immunoreactive oligodendroglia. It was suggested that some type of oligodendrocytes, specially those in perineuronal location, might fulfill a functional role more akin to astrocytes than to the normally myelinating oligodendroglia.

  9. What is the oligoadenylate synthetases-like protein and does it have therapeutic potential for influenza?

    PubMed Central

    Alcorn, John F.; Sarkar, Saumendra N.

    2015-01-01

    Besides its pandemic potential, seasonal influenza infection is associated with an estimated 250,000 to 500,000 deaths worldwide every year. Part of this virulence of influenza virus can be attributed to its ability to evade the host innate immune response. Here we discuss the possibility of using a recently described mechanism of boosting the innate immunity by oligoadenylate synthetase-like protein, to combat influenza infections. PMID:25544107

  10. Reassimilation of Photorespiratory Ammonium in Lotus japonicus Plants Deficient in Plastidic Glutamine Synthetase

    PubMed Central

    Pérez-Delgado, Carmen M.; García-Calderón, Margarita; Márquez, Antonio J.; Betti, Marco

    2015-01-01

    It is well established that the plastidic isoform of glutamine synthetase (GS2) is the enzyme in charge of photorespiratory ammonium reassimilation in plants. The metabolic events associated to photorespiratory NH4+ accumulation were analyzed in a Lotus japonicus photorespiratory mutant lacking GS2. The mutant plants accumulated high levels of NH4+ when photorespiration was active, followed by a sudden drop in the levels of this compound. In this paper it was examined the possible existence of enzymatic pathways alternative to GS2 that could account for this decline in the photorespiratory ammonium. Induction of genes encoding for cytosolic glutamine synthetase (GS1), glutamate dehydrogenase (GDH) and asparagine synthetase (ASN) was observed in the mutant in correspondence with the diminishment of NH4+. Measurements of gene expression, polypeptide levels, enzyme activity and metabolite levels were carried out in leaf samples from WT and mutant plants after different periods of time under active photorespiratory conditions. In the case of asparagine synthetase it was not possible to determine enzyme activity and polypeptide content; however, an increased asparagine content in parallel with the induction of ASN gene expression was detected in the mutant plants. This increase in asparagine levels took place concomitantly with an increase in glutamine due to the induction of cytosolic GS1 in the mutant, thus revealing a major role of cytosolic GS1 in the reassimilation and detoxification of photorespiratory NH4+ when the plastidic GS2 isoform is lacking. Moreover, a diminishment in glutamate levels was observed, that may be explained by the induction of NAD(H)-dependent GDH activity. PMID:26091523

  11. [Isolation of tyrosyl-tRNA-synthetase from Thermus thermophilus HB-27].

    PubMed

    Iaremchuk, A D; Tukalo, M A; Egorova, S P; Konovalenko, A V; Matsuka, G Kh

    1990-01-01

    A method for isolating tyrosyl-tRNA synthetase from Thermus thermophilus is described, including ammonium sulfate fractionation, chromatography on DEAE-sepharose, hydroxyapatite, heparin-sepharose and hydrophobic chromatography on Toyopearl HW-65. The yield of the purified enzyme was 1.6 mg per 1 kg of T. thermophilus cells. The enzyme is a dimer protein of the alpha 2 type with molecular weight of 100 kDa.

  12. Geometric Attitude Controls And Estimations On The Special Orthogonal Group

    NASA Astrophysics Data System (ADS)

    Wu, Tse-Huai

    This dissertation is concerned with spacecraft attitude control and estimation problems from the point of view of geometric mechanics. The controllers and observers are built on the special orthogonal group without any parameterizations, where the attitude dynamics is treated in a global and unique manner. The dissertation is composed of three parts. A leader-follower attitude formation control scheme is reported such that the leader spacecraft control its absolute attitude with respect to the inertial reference frame and the follower spacecraft control relative attitude with respect to other spacecraft in the formation. The unique feature is that both the absolute attitude and the relative attitude control systems are developed directly in terms of the line-of-sight observations, where attitude determination and estimation processes are not required. Second, an angular velocity observer is developed such that the estimated angular velocity is guaranteed to converge to the true angular velocity asymptotically from almost all initial estimates. Then, the presented observer is integrated with a proportional-derivative attitude tracking controller to show a separation type property for attitude tracking in the absence of angular velocity measurements. A hybrid observer for the attitude dynamics of a rigid body is proposed to guarantee global asymptotic stability. By designing a set of attitude error functions, attitude estimates are expelled from undesired equilibria to achieve global asymptotic stability. To guarantee that the estimated attitudes evolve on the special orthogonal group, a numerical algorithm based on the Lie group method is presented.

  13. Orthogonal topography in the parallel input architecture of songbird HVC.

    PubMed

    Elliott, Kevin C; Wu, Wei; Bertram, Richard; Hyson, Richard L; Johnson, Frank

    2017-06-15

    Neural activity within the cortical premotor nucleus HVC (acronym is name) encodes the learned songs of adult male zebra finches (Taeniopygia guttata). HVC activity is driven and/or modulated by a group of five afferent nuclei (the Medial Magnocellular nucleus of the Anterior Nidopallium, MMAN; Nucleus Interface, NIf; nucleus Avalanche, Av; the Robust nucleus of the Arcopallium, RA; the Uvaeform nucleus, Uva). While earlier evidence suggested that HVC receives a uniformly distributed and nontopographic pattern of afferent input, recent evidence suggests this view is incorrect (Basista et al., ). Here, we used a double-labeling strategy (varying both the distance between and the axial orientation of dual tracer injections into HVC) to reveal a massively parallel and in some cases topographic pattern of afferent input. Afferent neurons target only one rostral or caudal location within medial or lateral HVC, and each HVC location receives convergent input from each afferent nucleus in parallel. Quantifying the distributions of single-labeled cells revealed an orthogonal topography in the organization of afferent input from MMAN and NIf, two cortical nuclei necessary for song learning. MMAN input is organized across the lateral-medial axis whereas NIf input is organized across the rostral-caudal axis. To the extent that HVC activity is influenced by afferent input during the learning, perception, or production of song, functional models of HVC activity may need revision to account for the parallel input architecture of HVC, along with the orthogonal input topography of MMAN and NIf. © 2017 Wiley Periodicals, Inc.

  14. Efficient and Adaptive Orthogonal Finite Element Representation of the Geopotential

    NASA Astrophysics Data System (ADS)

    Junkins, John L.; Younes, Ahmad Bani; Woollands, Robyn M.; Bai, Xiaoli

    2017-06-01

    We unify and extend classical results from function approximation theory and consider their utility in astrodynamics. Least square approximation, using the classical Chebyshev polynomials as basis functions, is reviewed for discrete samples of the to-be-approximated function. We extend the orthogonal approximation ideas to n-dimensions in a novel way, through the use of array algebra and Kronecker operations. Approximation of test functions illustrates the resulting algorithms and provides insight into the errors of approximation, as well as the associated errors arising when the approximations are differentiated or integrated. Two sets of applications are considered that are challenges in astrodynamics. The first application addresses local approximation of high degree and order geopotential models, replacing the global spherical harmonic series by a family of locally precise orthogonal polynomial approximations for efficient computation. A method is introduced which adapts the approximation degree radially, compatible with the truth that the highest degree approximations (to ensure maximum acceleration error < 10-9 m s-2, globally) are required near the Earth's surface, whereas lower degree approximations are required as radius increases. We show that a four order of magnitude speedup is feasible, with efficiency optimized using radial adaptation.

  15. Orthogonal Cherenkov sound in spin-orbit coupled systems.

    PubMed

    Smirnov, Sergey

    2015-06-17

    Conventionally the Cherenkov sound is governed by orbital degrees of freedom and is excited by supersonic particles. Additionally, it usually has a forward nature with a conic geometry known as the Cherenkov cone whose axis is oriented along the supersonic particle motion. Here we predict Cherenkov sound of a unique nature entirely resulting from the electronic spin degree of freedom and demonstrate a fundamentally distinct Cherenkov effect originating from essentially subsonic electrons in two-dimensional gases with both Bychkov-Rashba and Dresselhaus spin-orbit interactions. Specifically, we show that the axis of the conventional forward Cherenkov cone gets a nontrivial quarter-turn and at the same time the sound distribution strongly localizes around this rotated axis being now orthogonal to the subsonic particle motion. Apart from its fundamentally appealing nature, the orthogonal Cherenkov sound could have applications in planar semiconductor technology combining spin and acoustic phenomena to develop, e.g., acoustic amplifiers or sound sources with a flexible spin dependent orientation of the sound propagation.

  16. Impact Damage of 3D Orthogonal Woven Composite Circular Plates

    NASA Astrophysics Data System (ADS)

    Ji, Changgan; Sun, Baozhong; Qiu, Yiping; Gu, Bohong

    2007-11-01

    The damages of 3D orthogonal woven composite circular plate under quasi-static indentation and transverse impact were tested with Materials Test System (MTS) and modified split Hopkinson bar (SHPB) apparatus. The load vs. displacement curves during quasi-static penetration and impact were obtained to study the energy absorption of the composite plate. The fluctuation of the impact stress waves has been unveiled. Differences of the load-displacement curves between the quasi-static and impact loading are discussed. This work also aims at establishing a unit-cell model to analyze the damage of composites. A user material subroutine which named VUMAT for characterizing the constitutive relationship of the 3-D orthogonal woven composite and the damage evolution is incorporated with a finite element code ABAQUS/Explicit to simulate the impact damage process of the composite plates. From the comparison of the load-displacement curves and energy absorption curves of the composite plate between experimental and FEM simulation, it is shown that the unit-cell model of the 3D woven composite and the VUMAT combined with the ABAQUS/Explicit can calculate the impact responses of the circular plate precisely. Furthermore, the model can also be extended to simulate the impact behavior of the 3D woven composite structures.

  17. Supervised orthogonal discriminant subspace projects learning for face recognition.

    PubMed

    Chen, Yu; Xu, Xiao-Hong

    2014-02-01

    In this paper, a new linear dimension reduction method called supervised orthogonal discriminant subspace projection (SODSP) is proposed, which addresses high-dimensionality of data and the small sample size problem. More specifically, given a set of data points in the ambient space, a novel weight matrix that describes the relationship between the data points is first built. And in order to model the manifold structure, the class information is incorporated into the weight matrix. Based on the novel weight matrix, the local scatter matrix as well as non-local scatter matrix is defined such that the neighborhood structure can be preserved. In order to enhance the recognition ability, we impose an orthogonal constraint into a graph-based maximum margin analysis, seeking to find a projection that maximizes the difference, rather than the ratio between the non-local scatter and the local scatter. In this way, SODSP naturally avoids the singularity problem. Further, we develop an efficient and stable algorithm for implementing SODSP, especially, on high-dimensional data set. Moreover, the theoretical analysis shows that LPP is a special instance of SODSP by imposing some constraints. Experiments on the ORL, Yale, Extended Yale face database B and FERET face database are performed to test and evaluate the proposed algorithm. The results demonstrate the effectiveness of SODSP. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Orthogonalizing EM: A design-based least squares algorithm.

    PubMed

    Xiong, Shifeng; Dai, Bin; Huling, Jared; Qian, Peter Z G

    We introduce an efficient iterative algorithm, intended for various least squares problems, based on a design of experiments perspective. The algorithm, called orthogonalizing EM (OEM), works for ordinary least squares and can be easily extended to penalized least squares. The main idea of the procedure is to orthogonalize a design matrix by adding new rows and then solve the original problem by embedding the augmented design in a missing data framework. We establish several attractive theoretical properties concerning OEM. For the ordinary least squares with a singular regression matrix, an OEM sequence converges to the Moore-Penrose generalized inverse-based least squares estimator. For ordinary and penalized least squares with various penalties, it converges to a point having grouping coherence for fully aliased regression matrices. Convergence and the convergence rate of the algorithm are examined. Finally, we demonstrate that OEM is highly efficient for large-scale least squares and penalized least squares problems, and is considerably faster than competing methods when n is much larger than p. Supplementary materials for this article are available online.

  19. Orthogonalizing EM: A design-based least squares algorithm

    PubMed Central

    Xiong, Shifeng; Dai, Bin; Huling, Jared; Qian, Peter Z. G.

    2016-01-01

    We introduce an efficient iterative algorithm, intended for various least squares problems, based on a design of experiments perspective. The algorithm, called orthogonalizing EM (OEM), works for ordinary least squares and can be easily extended to penalized least squares. The main idea of the procedure is to orthogonalize a design matrix by adding new rows and then solve the original problem by embedding the augmented design in a missing data framework. We establish several attractive theoretical properties concerning OEM. For the ordinary least squares with a singular regression matrix, an OEM sequence converges to the Moore-Penrose generalized inverse-based least squares estimator. For ordinary and penalized least squares with various penalties, it converges to a point having grouping coherence for fully aliased regression matrices. Convergence and the convergence rate of the algorithm are examined. Finally, we demonstrate that OEM is highly efficient for large-scale least squares and penalized least squares problems, and is considerably faster than competing methods when n is much larger than p. Supplementary materials for this article are available online. PMID:27499558

  20. Texel-based image classification with orthogonal bases

    NASA Astrophysics Data System (ADS)

    Carbajal-Degante, Erik; Nava, Rodrigo; Olveres, Jimena; Escalante-Ramírez, Boris; Kybic, Jan

    2016-04-01

    Periodic variations in patterns within a group of pixels provide important information about the surface of interest and can be used to identify objects or regions. Hence, a proper analysis can be applied to extract particular features according to some specific image properties. Recently, texture analysis using orthogonal polynomials has gained attention since polynomials characterize the pseudo-periodic behavior of textures through the projection of the pattern of interest over a group of kernel functions. However, the maximum polynomial order is often linked to the size of the texture, which implies in many cases, a complex calculation and introduces instability in higher orders leading to computational errors. In this paper, we address this issue and explore a pre-processing stage to compute the optimal size of the window of analysis called "texel." We propose Haralick-based metrics to find the main oscillation period, such that, it represents the fundamental texture and captures the minimum information, which is sufficient for classification tasks. This procedure avoids the computation of large polynomials and reduces substantially the feature space with small classification errors. Our proposal is also compared against different fixed-size windows. We also show similarities between full-image representations and the ones based on texels in terms of visual structures and feature vectors using two different orthogonal bases: Tchebichef and Hermite polynomials. Finally, we assess the performance of the proposal using well-known texture databases found in the literature.

  1. Bifurcations in two-image photometric stereo for orthogonal illuminations

    NASA Astrophysics Data System (ADS)

    Kozera, R.; Prokopenya, A.; Noakes, L.; Śluzek, A.

    2017-07-01

    This paper discusses the ambiguous shape recovery in two-image photometric stereo for a Lambertian surface. The current uniqueness analysis refers to linearly independent light-source directions p = (0, 0, -1) and q arbitrary. For this case necessary and sufficient condition determining ambiguous reconstruction is governed by a second-order linear partial differential equation with constant coefficients. In contrast, a general position of both non-colinear illumination directions p and q leads to a highly non-linear PDE which raises a number of technical difficulties. As recently shown, the latter can also be handled for another family of orthogonal illuminations parallel to the OXZ-plane. For the special case of p = (0, 0, -1) a potential ambiguity stems also from the possible bifurcations of sub-local solutions glued together along a curve defined by an algebraic equation in terms of the data. This paper discusses the occurrence of similar bifurcations for such configurations of orthogonal light-source directions. The discussion to follow is supplemented with examples based on continuous reflectance map model and generated synthetic images.

  2. Tomographic Approach in Three-Orthogonal-Basis Quantum Key Distribution

    NASA Astrophysics Data System (ADS)

    Liang, Wen-Ye; Wen, Hao; Yin, Zhen-Qiang; Chen, Hua; Li, Hong-Wei; Chen, Wei; Han, Zheng-Fu

    2015-09-01

    At present, there is an increasing awareness of some three-orthogonal-basis quantum key distribution protocols, such as, the reference-frame-independent (RFI) protocol and the six-state protocol. For secure key rate estimations of these protocols, there are two methods: one is the conventional approach, and another is the tomographic approach. However, a comparison between these two methods has not been given yet. In this work, with the general model of rotation channel, we estimate the key rate using conventional and tomographic methods respectively. Results show that conventional estimation approach in RFI protocol is equivalent to tomographic approach only in the case of that one of three orthogonal bases is always aligned. In other cases, tomographic approach performs much better than the respective conventional approaches of the RFI protocol and the six-state protocol. Furthermore, based on the experimental data, we illustrate the deep connections between tomography and conventional RFI approach representations. Supported by the National Basic Research Program of China under Grant Nos. 2011CBA00200 and 2011CB921200 and the National Natural Science Foundation of China under Grant Nos. 60921091, 61475148, and 61201239 and Zhejiang Natural Science Foundation under Grant No. LQ13F050005

  3. Langevin dynamics of a heavy particle and orthogonality effects

    NASA Astrophysics Data System (ADS)

    Thomas, Mark; Karzig, Torsten; Viola Kusminskiy, Silvia

    2015-12-01

    The dynamics of a classical heavy particle moving in a quantum environment is determined by a Langevin equation which encapsulates the effect of the environment-induced reaction forces on the particle. For an open quantum system, these include a Born-Oppenheimer force, a dissipative force, and a stochastic force due to shot and thermal noise. Recently, it was shown that these forces can be expressed in terms of the scattering matrix of the system by considering the classical heavy particle as a time-dependent scattering center, allowing to demonstrate interesting features of these forces when the system is driven out of equilibrium. At the same time, it is well known that small changes in a scattering potential can have a profound impact on a fermionic system due to the Anderson orthogonality catastrophe. In this work, by calculating the Loschmidt echo, we relate Anderson orthogonality effects with the mesoscopic reaction forces for an environment that can be taken out of equilibrium. In particular, we show how the decay of the Loschmidt echo is characterized by fluctuations and dissipation in the system and discuss different quench protocols.

  4. Cerenkov luminescence tomography based on preconditioning orthogonal matching pursuit

    NASA Astrophysics Data System (ADS)

    Liu, Haixiao; Hu, Zhenhua; Wang, Kun; Tian, Jie; Yang, Xin

    2015-03-01

    Cerenkov luminescence imaging (CLI) is a novel optical imaging method and has been proved to be a potential substitute of the traditional radionuclide imaging such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT). This imaging method inherits the high sensitivity of nuclear medicine and low cost of optical molecular imaging. To obtain the depth information of the radioactive isotope, Cerenkov luminescence tomography (CLT) is established and the 3D distribution of the isotope is reconstructed. However, because of the strong absorption and scatter, the reconstruction of the CLT sources is always converted to an ill-posed linear system which is hard to be solved. In this work, the sparse nature of the light source was taken into account and the preconditioning orthogonal matching pursuit (POMP) method was established to effectively reduce the ill-posedness and obtain better reconstruction accuracy. To prove the accuracy and speed of this algorithm, a heterogeneous numerical phantom experiment and an in vivo mouse experiment were conducted. Both the simulation result and the mouse experiment showed that our reconstruction method can provide more accurate reconstruction result compared with the traditional Tikhonov regularization method and the ordinary orthogonal matching pursuit (OMP) method. Our reconstruction method will provide technical support for the biological application for Cerenkov luminescence.

  5. Statistical benchmarking for orthogonal electrostatic quantum dot qubit devices

    NASA Astrophysics Data System (ADS)

    Gamble, John; Frees, Adam; Friesen, Mark; Coppersmith, S. N.

    2014-03-01

    Quantum dots in semiconductor systems have emerged as attractive candidates for the implementation of quantum information processors because of the promise of scalability, manipulability, and integration with existing classical electronics. A limitation in current devices is that the electrostatic gates used for qubit manipulation exhibit strong cross-capacitance, presenting a barrier for practical scale-up. Here, we introduce a statistical framework for making precise the notion of orthogonality. We apply our method to analyze recently implemented designs at the University of Wisconsin-Madison that exhibit much increased orthogonal control than was previously possible. We then use our statistical modeling to future device designs, providing practical guidelines for devices to have robust control properties. Sandia National Laboratories is a multi-program laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin Corporation, for the U.S. Department of Energy Nuclear Security Administration under contract DE-AC04-94AL85000. The views and conclusions contained in this document are those of the authors and should not be interpreted as representing the official policies, either expressly or implied, of the US Government. This work was supported in part by the Laboratory Directed Research and Development program at Sandia National Laboratories, by ARO (W911NF-12-0607), and by the United States Department of Defense.

  6. Structural Basis for Specific Inhibition of tRNA Synthetase by an ATP Competitive Inhibitor.

    PubMed

    Fang, Pengfei; Han, Hongyan; Wang, Jing; Chen, Kaige; Chen, Xin; Guo, Min

    2015-06-18

    Pharmaceutical inhibitors of aminoacyl-tRNA synthetases demand high species and family specificity. The antimalarial ATP-mimetic cladosporin selectively inhibits Plasmodium falciparum LysRS (PfLysRS). How the binding to a universal ATP site achieves the specificity is unknown. Here we report three crystal structures of cladosporin with human LysRS, PfLysRS, and a Pf-like human LysRS mutant. In all three structures, cladosporin occupies the class defining ATP-binding pocket, replacing the adenosine portion of ATP. Three residues holding the methyltetrahydropyran moiety of cladosporin are critical for the specificity of cladosporin against LysRS over other class II tRNA synthetase families. The species-exclusive inhibition of PfLysRS is linked to a structural divergence beyond the active site that mounts a lysine-specific stabilizing response to binding cladosporin. These analyses reveal that inherent divergence of tRNA synthetase structural assembly may allow for highly specific inhibition even through the otherwise universal substrate binding pocket and highlight the potential for structure-driven drug development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Food safety: Structure and expression of the asparagine synthetase gene family of wheat

    PubMed Central

    Gao, Runhong; Curtis, Tanya Y.; Powers, Stephen J.; Xu, Hongwei; Huang, Jianhua; Halford, Nigel G.

    2016-01-01

    Asparagine is an important nitrogen storage and transport molecule, but its accumulation as a free amino acid in crops has implications for food safety because free asparagine is a precursor for acrylamide formation during cooking and processing. Asparagine synthesis occurs by the amidation of aspartate, catalysed by asparagine synthetase, and this study concerned the expression of asparagine synthetase (TaASN) genes in wheat. The expression of three genes, TaASN1-3, was studied in different tissues and in response to nitrogen and sulphur supply. The expression of TaASN2 in the embryo and endosperm during mid to late grain development was the highest of any of the genes in any tissue. Both TaASN1 and TaASN2 increased in expression through grain development, and in the grain of field-grown plants during mid-development in response to sulphur deprivation. However, only TaASN1 was affected by nitrogen or sulphur supply in pot-based experiments, showing complex tissue-specific and developmentally-changing responses. A putative N-motif or GCN4-like regulatory motif was found in the promoter of TaASN1 genes from several cereal species. As the study was completed, a fourth gene, TaASN4, was identified from recently available genome data. Phylogenetic analysis showed that other cereal species have similar asparagine synthetase gene families to wheat. PMID:27110058

  8. Food safety: Structure and expression of the asparagine synthetase gene family of wheat.

    PubMed

    Gao, Runhong; Curtis, Tanya Y; Powers, Stephen J; Xu, Hongwei; Huang, Jianhua; Halford, Nigel G

    2016-03-01

    Asparagine is an important nitrogen storage and transport molecule, but its accumulation as a free amino acid in crops has implications for food safety because free asparagine is a precursor for acrylamide formation during cooking and processing. Asparagine synthesis occurs by the amidation of aspartate, catalysed by asparagine synthetase, and this study concerned the expression of asparagine synthetase (TaASN) genes in wheat. The expression of three genes, TaASN1-3, was studied in different tissues and in response to nitrogen and sulphur supply. The expression of TaASN2 in the embryo and endosperm during mid to late grain development was the highest of any of the genes in any tissue. Both TaASN1 and TaASN2 increased in expression through grain development, and in the grain of field-grown plants during mid-development in response to sulphur deprivation. However, only TaASN1 was affected by nitrogen or sulphur supply in pot-based experiments, showing complex tissue-specific and developmentally-changing responses. A putative N-motif or GCN4-like regulatory motif was found in the promoter of TaASN1 genes from several cereal species. As the study was completed, a fourth gene, TaASN4, was identified from recently available genome data. Phylogenetic analysis showed that other cereal species have similar asparagine synthetase gene families to wheat.

  9. Oxidative modification of glutamine synthetase. I. Inactivation is due to loss of one histidine residue.

    PubMed

    Levine, R L

    1983-10-10

    Intracellular proteolytic degradation of glutamine synthetase occurs in two distinct steps in Escherichia coli (Levine, R. L., Oliver, C. N., Fulks, R. M., and Stadtman, E. R. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 2120-2124). In the first step, a mixed function oxidation modifies the glutamine synthetase. The modified enzyme, which is catalytically inactive, becomes susceptible to proteolytic attack. In the second step, a protease specific for the modified enzyme catalyzes the actual proteolytic degradation. The oxidatively modified glutamine synthetase was studied to determine the chemical differences between it and the native enzyme. Only a single alteration was found; one of sixteen histidine residues/subunit was altered by the oxidative modification. The modification introduced a carbonyl group into the protein, permitting isolation of a stable dinitrophenylhydrazone. No other differences were detected between the native and modified proteins. Specifically, the cysteine, methionine, phenylalanine, tyrosine, and tryptophan contents were not altered. A number of other prokaryotic and eukaryotic enzymes are also susceptible to oxidative modification. This covalent modification may be important in intracellular proteolysis, in mammalian host defense systems, in prevention of autolysis, in aging processes, and in oxygen toxicity.

  10. Dual binding sites for translocation catalysis by Escherichia coli glutathionylspermidine synthetase.

    PubMed

    Pai, Chien-Hua; Chiang, Bing-Yu; Ko, Tzu-Ping; Chou, Chia-Cheng; Chong, Cheong-Meng; Yen, Fang-Jiun; Chen, Shoujun; Coward, James K; Wang, Andrew H-J; Lin, Chun-Hung

    2006-12-13

    Most organisms use glutathione to regulate intracellular thiol redox balance and protect against oxidative stress; protozoa, however, utilize trypanothione for this purpose. Trypanothione biosynthesis requires ATP-dependent conjugation of glutathione (GSH) to the two terminal amino groups of spermidine by glutathionylspermidine synthetase (GspS) and trypanothione synthetase (TryS), which are considered as drug targets. GspS catalyzes the penultimate step of the biosynthesis-amide bond formation between spermidine and the glycine carboxylate of GSH. We report herein five crystal structures of Escherichia coli GspS in complex with substrate, product or inhibitor. The C-terminal of GspS belongs to the ATP-grasp superfamily with a similar fold to the human glutathione synthetase. GSH is likely phosphorylated at one of two GSH-binding sites to form an acylphosphate intermediate that then translocates to the other site for subsequent nucleophilic addition of spermidine. We also identify essential amino acids involved in the catalysis. Our results constitute the first structural information on the biochemical features of parasite homologs (including TryS) that underlie their broad specificity for polyamines.

  11. Brucella melitensis Methionyl-tRNA-Synthetase (MetRS), a Potential Drug Target for Brucellosis

    PubMed Central

    Ranade, Ranae M.; Zhang, Zhongsheng; Dranow, David M.; Myers, Janette B.; Choi, Ryan; Nakazawa Hewitt, Steve; Edwards, Thomas E.; Davies, Douglas R.; Lorimer, Donald; Boyle, Stephen M.; Barrett, Lynn K.; Buckner, Frederick S.; Fan, Erkang; Van Voorhis, Wesley C.

    2016-01-01

    We investigated Brucella melitensis methionyl-tRNA-synthetase (BmMetRS) with molecular, structural and phenotypic methods to learn if BmMetRS is a promising target for brucellosis drug development. Recombinant BmMetRS was expressed, purified from wild type Brucella melitensis biovar Abortus 2308 strain ATCC/CRP #DD-156 and screened by a thermal melt assay against a focused library of one hundred previously classified methionyl-tRNA-synthetase inhibitors of the blood stage form of Trypanosoma brucei. Three compounds showed appreciable shift of denaturation temperature and were selected for further studies on inhibition of the recombinant enzyme activity and cell viability against wild type B. melitensis strain 16M. BmMetRS protein complexed with these three inhibitors resolved into three-dimensional crystal structures and was analyzed. All three selected methionyl-tRNA-synthetase compounds inhibit recombinant BmMetRS enzymatic functions in an aminoacylation assay at varying concentrations. Furthermore, growth inhibition of B. melitensis strain 16M by the compounds was shown. Inhibitor-BmMetRS crystal structure models were used to illustrate the molecular basis of the enzyme inhibition. Our current data suggests that BmMetRS is a promising target for brucellosis drug development. However, further studies are needed to optimize lead compound potency, efficacy and safety as well as determine the pharmacokinetics, optimal dosage, and duration for effective treatment. PMID:27500735

  12. Brucella melitensis Methionyl-tRNA-Synthetase (MetRS), a Potential Drug Target for Brucellosis.

    PubMed

    Ojo, Kayode K; Ranade, Ranae M; Zhang, Zhongsheng; Dranow, David M; Myers, Janette B; Choi, Ryan; Nakazawa Hewitt, Steve; Edwards, Thomas E; Davies, Douglas R; Lorimer, Donald; Boyle, Stephen M; Barrett, Lynn K; Buckner, Frederick S; Fan, Erkang; Van Voorhis, Wesley C

    2016-01-01

    We investigated Brucella melitensis methionyl-tRNA-synthetase (BmMetRS) with molecular, structural and phenotypic methods to learn if BmMetRS is a promising target for brucellosis drug development. Recombinant BmMetRS was expressed, purified from wild type Brucella melitensis biovar Abortus 2308 strain ATCC/CRP #DD-156 and screened by a thermal melt assay against a focused library of one hundred previously classified methionyl-tRNA-synthetase inhibitors of the blood stage form of Trypanosoma brucei. Three compounds showed appreciable shift of denaturation temperature and were selected for further studies on inhibition of the recombinant enzyme activity and cell viability against wild type B. melitensis strain 16M. BmMetRS protein complexed with these three inhibitors resolved into three-dimensional crystal structures and was analyzed. All three selected methionyl-tRNA-synthetase compounds inhibit recombinant BmMetRS enzymatic functions in an aminoacylation assay at varying concentrations. Furthermore, growth inhibition of B. melitensis strain 16M by the compounds was shown. Inhibitor-BmMetRS crystal structure models were used to illustrate the molecular basis of the enzyme inhibition. Our current data suggests that BmMetRS is a promising target for brucellosis drug development. However, further studies are needed to optimize lead compound potency, efficacy and safety as well as determine the pharmacokinetics, optimal dosage, and duration for effective treatment.

  13. Lactose synthetase activity in mouse mammary glands is controlled by thyroid hormones

    PubMed Central

    1979-01-01

    Epithelial cells in explants from the mammary glands of euthyroid mature virgin mice are proliferatively dormant. They must undergo DNA synthesis and traverse the cell cycle in vitro before they are able to differentiate fully in response to insulin, hydrocortisone, and prolactin, and synthesize enzymatically active alpha-lactalbumin (measured as lactose synthetase activity). In contrast, glands from hyperthyroid mature virgin mice do not require DNA synthesis in vitro to differentiate. Explants from the euthyroid virgin tissue overcome their dependence on DNA synthesis when 10(-9) M 3,5,3'-triiodo-L- thyronine is added directly to the cultures in addition to the other three hormones. Explants from involuted mammary glands from euthyroid primiparous mice do not require DNA synthesis in vitro to make the milk protein even though they, like explants from mature euthyroid virgin tissue, are proliferatively dormant and do not contain detectable lactose synthetase activity in vivo. Glands from primiparous animals made mildly hypothyroid by ingestion of 0.1% thiouracil in drinking water during 7 wk of involution remain morphologically indistinguishable from glands of their euthyroid counterparts. However, explants from the glands of these hypothyroid animals revert to a state of dependence on DNA synthesis to differentiate functionally. These observations suggest that the dependence on DNA synthesis and cell cycle traversal for hormonal induction of lactose synthetase activity in the mouse mammary gland is controlled by thyroid hormones. PMID:117014

  14. Beneficial consequences of a selective glutamine synthetase inhibitor in oats and legumes

    SciTech Connect

    Langston-Unkefer, P.J.; Knight, T.J.; Sengupta-Gopalan, C.

    1988-01-01

    We report on the effects of administering a unique glutamine synthetase inhibitor to cereals and N/sub 2/-fixing legumes. A bacterium (Pseudomonas syringae pv. tabaci) delivers this inhibitor to provide extended treatment periods; we inoculated the root systems of oat and legume plants with pv. tabaci to provide for delivery of this inhibitor to their root or root/nodule systems. Inoculation of legumes is accompanied by increased plant growth, total plant nitrogen, nodulation, and nitrogen fixation activity. Inoculation of the oats is accompanied by either of two results depending upon the genotype of the oat plant. One result is inhibition of plant growth followed by plant death as consequences of the loss of all of the glutamine synthetase activities in the plant and the subsequent accumulation of ammonia and cessation of nitrate uptake. The second and opposite result is observed in a small population of oats screened from a commercial cultivar and includes increased plant growth and leaf protein. The effects of this inhibitor can be beneficial when applied to appropriate plant material. In an attempt to effectively communicate these findings to the reader, we first introduce the inhibitor (a novel amino acid) and its bacterial delivery systems, the target of the inhibitor (glutamine synthetase-catalyzed ammonia assimilation), and the two different nitrogen economics in the legume and cereal plants used experimentally. The physiological, biochemical, and molecular genetic consequences of the inhibitor action in cereals and legumes, as we presently understand them, are then presented. 18 refs., 4 figs., 3 tabs.,

  15. Directed evolution of adenylosuccinate synthetase from Bacillus subtilis and its application in metabolic engineering.

    PubMed

    Wang, Xiaoyue; Wang, Guanglu; Li, Xinli; Fu, Jing; Chen, Tao; Wang, Zhiwen; Zhao, Xueming

    2016-08-10

    Adenylosuccinate synthetase (EC. 6.3.4.4) encoded by purA in Bacillus subtilis, catalyzing the first step of the conversion of IMP to AMP, plays an important role in flux distribution in the purine biosynthetic pathway. In this study, we described the use of site saturation mutagenesis to obtain a desired enzyme activity of adenylosuccinate synthetase and its application in flux regulation. Based on sequence alignment and structural modeling, a library of enzyme variants was created by a semi-rational evolution strategy in position Thr238 and Pro242. Other than purA deletion, the leaky mutation purA(P242N) partially reduced the flux towards AMP derived from IMP and increased the riboflavin synthesis precursor GTP, while also kept the requirement of ATP synthesis for cell growth. PurA(P242N) was introduced into an inosine-producing strain and resulted in an approximately 4.66-fold increase in inosine production, from 0.088±0.009g/L to 0.41±0.051g/L, in minimal medium without hypoxanthine accumulation. These results underline that the directed evolution of adenylosuccinate synthetase could tailor its activities and adjust metabolic flux. This mutation may provide a promising application in purine-based product accumulation, like inosine, guanosine and folate which are directly stemming from purine pathway in B. subtilis.

  16. Comparison of effects of aspirin and indomethacin on human platelet prostaglandin synthetase.

    PubMed Central

    Crook, D; Collins, A J

    1977-01-01

    Human platelets were incubated in vitro with either aspirin or indomethacin and the prostaglandin synthetase activity of the resultant microsomal fraction from each incubation measured using a radiometric technique. Whereas aspirin produced a dose-related inhibition of the enzyme, indomethacin produced little or no inhibition over the same concentration range (10(-6) mol/l--10(-3) mol/l). Furthermore, administration of aspirin (600 mg) to volunteers produced a highly significant, prolonged inhibition of platelet microsomal prostaglandin synthetase whereas no inhibition was found with indomethacin (50 mg). As indomethacin is considerably more potent than aspirin as an inhibitor of human platelet prostaglandin synthetase in vitro, the results suggest a fundamental difference in the nature of the inhibition produced by each drug, aspirin being an essentially irreversible inhibitor whereas the inhibition produced by indomethacin is reversible. Studies with [3H-acetyl] aspirin have confirmed previous findings (Roth and Majerus, 1975) that aspirin produces an irreversible acetylation of a particulate fraction protein from human platelets. PMID:411427

  17. Regulation of 2', 5'-oligoadenylate synthetase gene expression by interferons and platelet-derived growth factor

    SciTech Connect

    Garcia-Blanco, M.A. ); Lengyel, P. . Dept. of Molecular Biophysics and Biochemistry); Morrison, E.; BrownLee, C.; Stiles, C.D. ); Williams, B.R.G. )

    1989-03-01

    In murine BALB/c 3T3 cell cultures, either beta interferon or platelet-derived growth factor (PDGF) enhanced expression of the 2', 5-oligoadenylate synthetase mRNA and protein. The time course of induction in response to beta inteferon was similar to that in response to PDGF. Of several growth factors known to be present in clotted blood serum (i.e., epidermal growth factor, transforming growth factor beta, and PDGF), only PDGF enhanced expression of 2', 5-oligoadenylate synthetase. The linkage of an interferon response element-containing segment from the 5'-flanking region of a human or murine 2'-5'-oligoadenylate synthetase gene made a heterologous gene responsive to interferon. The expression of such a gene construct in transfected cells was also induced by PDGF. Induction by PDGF was inhibited by mono- or polyclonal antibodies to murine interferon, which suggested that induction by PDGF requires interferon. Both PDGF and interferon induced nuclear factors that bound to this interferon response element-containing segment in vitro.

  18. Gene organization around the phenylalanyl-transfer ribonucleic acid synthetase locus in Escherichia coli.

    PubMed Central

    Comer, M M

    1981-01-01

    The organization of seven genes located at about 38 min on the genetic map of Escherichia coli was examined; these genes included pheS and pheT, which code for the alpha and beta subunits of phenylalanyl-transfer ribonucleic acid synthetase, and thrS, the structural gene for threonyl-transfer ribonucleic acid synthetase. Deletion mutants were isolated from an F-prime-containing merodiploid strain and were characterized genetically. Seventeen different kinds of deletions extending into pheS of pheT were identified. These deletions unambiguously defined the gene order as aroD pps himA pheT pheS thrS pfkB. Mutants with deletions covering either pheS or pheT, but not both, were analyzed further by assay of phenylalanyl-transfer ribonucleic acid synthetase. The phenotype of the mutants with a deletion from pfkB through pheS was anomalous; although the pheT gene was apparently still present, its product, the beta subunit, was much reduced in activity. PMID:7012115

  19. Cloning and functional characterization of a homoglutathione synthetase from pea nodules.

    PubMed

    Iturbe-Ormaetxe, Iñaki; Heras, Begoña; Matamoros, Manuel A; Ramos, Javier; Moran, Jose F; Becana, Manuel

    2002-05-01

    The thiol tripeptide glutathione (GSH; gammaGlu-Cys-Gly) is very abundant in legume nodules where it performs multiple functions that are critical for optimal nitrogen fixation. Some legume nodules contain another tripeptide, homoglutathione (hGSH; gammaGlu-Cys-betaAla), in addition to or instead of GSH. We have isolated from a pea (Pisum sativum L.) nodule library a cDNA, GSHS2, that is expressed in nodules but not in leaves. This cDNA was overexpressed in insect cells and its protein product was identified as a highly active and specific hGSH synthetase. The enzyme, the first of this type to be completely purified, is predicted to be a homodimeric cytosolic protein. It shows a specific activity of 3400 nmol hGSH min-1 mg-1 protein with a standard substrate concentration (5 mM beta-alanine) and Km values of 1.9 mM for beta-alanine and 104 mM for glycine. The specificity constant (Vmax/Km) shows that the pure enzyme is 57.3-fold more specific for beta-alanine than for glycine. Southern blot analysis revealed that the gene is present as a single copy in the pea genome and that there are homologous genes in other legumes. We conclude that the synthesis of hGSH in pea nodules is catalysed by a specific hGSH synthetase and not by a GSH synthetase with broad substrate specificity.

  20. Unique domain appended to vertebrate tRNA synthetase is essential for vascular development

    PubMed Central

    Xu, Xiaoling; Shi, Yi; Zhang, Hui-Min; Swindell, Eric C.; Marshall, Alan G.; Guo, Min; Kishi, Shuji; Yang, Xiang-Lei

    2012-01-01

    New domains were progressively added to cytoplasmic aminoacyl transfer RNA (tRNA) synthetases during evolution. One example is the UNE-S domain, appended to seryl-tRNA synthetase (SerRS) in species that developed closed circulatory systems. Here we show using solution and crystal structure analyses and in vitro and in vivo functional studies that UNE-S harbours a robust nuclear localization signal (NLS) directing SerRS to the nucleus where it attenuates vascular endothelial growth factor A expression. We also show that SerRS mutants previously linked to vasculature abnormalities either deleted the NLS or have the NLS sequestered in an alternative conformation. A structure-based second-site mutation, designed to release the sequestered NLS, restored normal vasculature. Thus, the essential function of SerRS in vascular development depends on UNE-S. These results are the first to show an essential role for a tRNA synthetase-associated appended domain at the organism level, and suggest that acquisition of UNE-S has a role in the establishment of the closed circulatory systems of vertebrates. PMID:22353712