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Sample records for orthogonal trna-aminoacyl-trna synthetase

  1. 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

    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.

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

    DOEpatents

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

    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 composition for the production of orthogonal tRNA-aminoacyltRNA synthetase pairs

    DOEpatents

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

    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.

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

    DOEpatents

    Schultz, Peter G.; 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

    2010-05-11

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

    DOEpatents

    Anderson, J Christopher [San Francisco, CA; Schultz, Peter G [La Jolla, CA; Santoro, Stephen [Cambridge, MA

    2009-05-05

    Compositions and methods of producing components of protein biosynthetic machinery that include glutamyl orthogonal tRNAs, glutamyl orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of glutamyl tRNAs/synthetases are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins using these orthogonal pairs.

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

    SciTech Connect

    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.

  7. Methods and composition 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

    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.

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

    DOEpatents

    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.

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

    DOEpatents

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

    2009-08-18

    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.

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

    DOEpatents

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

    2009-12-29

    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.

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

    DOEpatents

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

    2011-10-04

    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.

  12. 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.

  13. 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

  14. 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.

  15. 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.

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

    SciTech Connect

    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.

  17. Orthogonal polynomials and tolerancing

    NASA Astrophysics Data System (ADS)

    Rogers, John R.

    2011-10-01

    Previous papers have established the inadvisability of applying tolerances directly to power-series aspheric coefficients. The basic reason is that the individual terms are far from orthogonal. Zernike surfaces and the new Forbes surface types have certain orthogonality properties over the circle described by the "normalization radius." However, at surfaces away from the stop, the optical beam is smaller than the surface, and the polynomials are not orthogonal over the area sampled by the beam. In this paper, we investigate the breakdown of orthogonality as the surface moves away from the aperture stop, and the implications of this to tolerancing.

  18. Orthogonal Regression and Equivariance.

    ERIC Educational Resources Information Center

    Blankmeyer, Eric

    Ordinary least-squares regression treats the variables asymmetrically, designating a dependent variable and one or more independent variables. When it is not obvious how to make this distinction, a researcher may prefer to use orthogonal regression, which treats the variables symmetrically. However, the usual procedure for orthogonal regression is…

  19. Orthogonal tensor decompositions

    SciTech Connect

    Tamara G. Kolda

    2000-03-01

    The authors explore the orthogonal decomposition of tensors (also known as multi-dimensional arrays or n-way arrays) using two different definitions of orthogonality. They present numerous examples to illustrate the difficulties in understanding such decompositions. They conclude with a counterexample to a tensor extension of the Eckart-Young SVD approximation theorem by Leibovici and Sabatier [Linear Algebra Appl. 269(1998):307--329].

  20. 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

  1. Reengineering orthogonally selective riboswitches.

    PubMed

    Dixon, Neil; Duncan, John N; Geerlings, Torsten; Dunstan, Mark S; McCarthy, John E G; Leys, David; Micklefield, Jason

    2010-02-16

    The ability to independently control the expression of multiple genes by addition of distinct small-molecule modulators has many applications from synthetic biology, functional genomics, pharmaceutical target validation, through to gene therapy. Riboswitches are relatively simple, small-molecule-dependent, protein-free, mRNA genetic switches that are attractive targets for reengineering in this context. Using a combination of chemical genetics and genetic selection, we have developed riboswitches that are selective for synthetic "nonnatural" small molecules and no longer respond to the natural intracellular ligands. The orthogonal selectivity of the riboswitches is also demonstrated in vitro using isothermal titration calorimetry and x-ray crystallography. The riboswitches allow highly responsive, dose-dependent, orthogonally selective, and dynamic control of gene expression in vivo. It is possible that this approach may be further developed to reengineer other natural riboswitches for application as small-molecule responsive genetic switches in both prokaryotes and eukaryotes. PMID:20133756

  2. 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.

  3. Neurospora crassa mutants deficient in asparagine synthetase.

    PubMed Central

    MacPhee, K G; Nelson, R E; Schuster, S M

    1983-01-01

    Neurospora crassa mutants deficient in asparagine synthetase were selected by using the procedure of inositol-less death. Complementation tests among the 100 mutants isolated suggested that their alterations were genetically allelic. Recombination analysis with strain S1007t, an asparagine auxotroph, indicated that the mutations were located near or within the asn gene on linkage group V. In vitro assays with a heterokaryon indicated that the mutation was dominant. Thermal instability of cell extracts from temperature-sensitive strains in an in vitro asparagine synthetase assay determined that the mutations were in the structural gene(s) for asparagine synthetase. PMID:6137480

  4. 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.

  5. Some discrete multiple orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Arvesú, J.; Coussement, J.; van Assche, W.

    2003-04-01

    In this paper, we extend the theory of discrete orthogonal polynomials (on a linear lattice) to polynomials satisfying orthogonality conditions with respect to r positive discrete measures. First we recall the known results of the classical orthogonal polynomials of Charlier, Meixner, Kravchuk and Hahn (T.S. Chihara, An Introduction to Orthogonal Polynomials, Gordon and Breach, New York, 1978; R. Koekoek and R.F. Swarttouw, Reports of the Faculty of Technical Mathematics and Informatics No. 98-17, Delft, 1998; A.F. Nikiforov et al., Classical Orthogonal Polynomials of a Discrete Variable, Springer, Berlin, 1991). These polynomials have a lowering and raising operator, which give rise to a Rodrigues formula, a second order difference equation, and an explicit expression from which the coefficients of the three-term recurrence relation can be obtained. Then we consider r positive discrete measures and define two types of multiple orthogonal polynomials. The continuous case (Jacobi, Laguerre, Hermite, etc.) was studied by Van Assche and Coussement (J. Comput. Appl. Math. 127 (2001) 317-347) and Aptekarev et al. (Multiple orthogonal polynomials for classical weights, manuscript). The families of multiple orthogonal polynomials (of type II) that we will study have a raising operator and hence a Rodrigues formula. This will give us an explicit formula for the polynomials. Finally, there also exists a recurrence relation of order r+1 for these multiple orthogonal polynomials of type II. We compute the coefficients of the recurrence relation explicitly when r=2.

  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. The Orthogonal Transfer CCD

    NASA Astrophysics Data System (ADS)

    Tonry, J.; Burke, Barry E.; Schechter, Paul L.

    1997-10-01

    We have designed and built a new type of CCD that we call an orthogonal transfer CCD (OTCCD), which permits parallel clocking horizontally as well as vertically. The device has been used successfully to remove image motion caused by atmospheric turbulence at rates up to 100 Hz, and promises to be a better, cheaper way to carry out image motion correction for imaging than by using fast tip/tilt mirrors. We report on the device characteristics, and find that the large number of transfers needed to track image motion does not significantly degrade the image either because of charge transfer inefficiency or because of charge traps. For example, after 100 sec of tracking at 100 Hz approximately 3% of the charge would diffuse into a skirt around the point spread function. Four nights of data at the Michigan-Dartmouth-MIT (MDM) 2.4-m telescope also indicate that the atmosphere is surprisingly benign, in terms of both the speed and coherence angle of image motion. Image motion compensation improved image sharpness by about 0.5'' in quadrature with no degradation over a field of at least 3 arcminutes. (SECTION: Astronomical Instrumentation)

  8. Molecular characterization of N-acetylaspartylglutamate synthetase.

    PubMed

    Becker, Ivonne; Lodder, Julia; Gieselmann, Volkmar; Eckhardt, Matthias

    2010-09-17

    The dipeptide N-acetylaspartyl-glutamate (NAAG) is an abundant neuropeptide in the mammalian brain. Despite this fact, its physiological role is poorly understood. NAAG is synthesized by a NAAG synthetase catalyzing the ATP-dependent condensation of N-acetylaspartate and glutamate. In vitro NAAG synthetase activity has not been described, and the enzyme has not been purified. Using a bioinformatics approach we identified a putative dipeptide synthetase specifically expressed in the nervous system. Expression of the gene, which we named NAAGS (for NAAG synthetase) was sufficient to induce NAAG synthesis in primary astrocytes or CHO-K1 and HEK-293T cells when they coexpressed the NAA transporter NaDC3. Furthermore, coexpression of NAAGS and the recently identified N-acetylaspartate (NAA) synthase, Nat8l, in CHO-K1 or HEK-293T cells was sufficient to enable these cells to synthesize NAAG. Identity of the reaction product of NAAGS was confirmed by HPLC and electrospray ionization tandem mass spectrometry (ESI-MS). High expression levels of NAAGS were restricted to the brain, spinal cord, and testis. Taken together our results strongly suggest that the identified gene encodes a NAAG synthetase. Its identification will enable further studies to examine the role of this abundant neuropeptide in the vertebrate nervous system. PMID:20643647

  9. 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.

  10. 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.

  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. Alternative splicing creates two new architectures for human tyrosyl-tRNA synthetase.

    PubMed

    Wei, Zhiyi; Xu, Zhiwen; Liu, Xiaotian; Lo, Wing-Sze; Ye, Fei; Lau, Ching-Fun; Wang, Feng; Zhou, Jie J; Nangle, Leslie A; Yang, Xiang-Lei; Zhang, Mingjie; Schimmel, Paul

    2016-02-18

    Many human tRNA synthetases evolved alternative functions outside of protein synthesis. These functions are associated with over 200 splice variants (SVs), most of which are catalytic nulls that engender new biology. While known to regulate non-translational activities, little is known about structures resulting from natural internal ablations of any protein. Here, we report analysis of two closely related, internally deleted, SVs of homodimeric human tyrosyl-tRNA synthetase (TyrRS). In spite of both variants ablating a portion of the catalytic core and dimer-interface contacts of native TyrRS, each folded into a distinct stable structure. Biochemical and nuclear magnetic resonance (NMR) analysis showed that the internal deletion of TyrRSΔE2-4 SV gave an alternative, neomorphic dimer interface 'orthogonal' to that of native TyrRS. In contrast, the internal C-terminal splice site of TyrRSΔE2-3 prevented either dimerization interface from forming, and yielded a predominantly monomeric protein. Unlike ubiquitous TyrRS, the neomorphs showed clear tissue preferences, which were distinct from each other. The results demonstrate a sophisticated structural plasticity of a human tRNA synthetase for architectural reorganizations that are preferentially elicited in specific tissues. PMID:26773056

  13. Nonlocality of orthogonal product states

    NASA Astrophysics Data System (ADS)

    Zhang, Zhi-Chao; Gao, Fei; Qin, Su-Juan; Yang, Ying-Hui; Wen, Qiao-Yan

    2015-07-01

    In this paper, we mainly study the local indistinguishability of mutually orthogonal product basis quantum states in d ⊗d . In 3 ⊗3 , Bennett et al. [ Phys. Rev. A 59, 1070 (1999), 10.1103/PhysRevA.59.1070] presented nine orthogonal product basis quantum states which cannot be distinguished by local operations and classical communication (LOCC). In the work by Zhang et al. [Z.-C. Zhang et al., Phys. Rev. A 90, 022313 (2014), 10.1103/PhysRevA.90.022313], this result was generalized in d ⊗d , where d is odd. In this paper, we aim to construct locally indistinguishable orthogonal product basis quantum states in d ⊗d . For the general d ⊗d (d >2 ) quantum system, we first construct 4 d -4 orthogonal product states, and prove these states are locally indistinguishable using a very simple but quite effective method. Then, based on these states, we construct some classes of locally indistinguishable orthogonal product basis quantum states (OPBS) in d ⊗d (d >2 ) . Finally, we construct some LOCC indistinguishable OPBS in multipartite quantum systems. All of the above results demonstrate the phenomenon of nonlocality without entanglement.

  14. Phosphorylation of eukaryotic aminoacyl-tRNA synthetases

    SciTech Connect

    Pendergast, A.M.

    1986-01-01

    The phosphorylation of the highly purified aminoacyl-tRNA synthetase complex from rabbit reticulocytes was examined. The synthetase complex contained, in addition to eight aminoacyl-tRNA synthetases, three unidentified proteins and was free of endogenous protein kinase activity. Incubation of the complex with casein kinase I in the presence of ATP resulted in the phosphorylation of four synthetases, the glutamyl-, isoleucyl-, methionyl-, and lysyl-tRNA synthetases. Phosphorylation by casein kinase I altered binding to tRNA-Sepharose such that the phosphorylated complex eluted at 190 mM NaCl instead of the 275 mM salt observed for the nonphosphorylated form. Phosphorylation by casein kinase I resulted in a significant inhibition of aminoacylation with the four synthetases; the activities of the nonphosphorylated synthetases were unchanged. One of the unidentified proteins in the complex (M/sub r/ 37,000) was also an excellent substrate for casein kinase I. A comparison of the properties and two-dimensional phosphopeptide pattern of this protein with that of casein kinase I suggest that the 37,000 dalton protein in the synthetase complex is an inactive form of casein kinase I. Two other protein kinases were shown to phosphorylate aminoacyl-tRNA synthetases in the complex. The phosphorylation of threonyl-tRNA synthetase was also investigated. Five aminoacyl-tRNA synthetases in the high molecular weight complex were shown to be phosphorylated in rabbit reticulocytes following labeling with (/sup 32/P)orthophosphate.

  15. Genetics Home Reference: glutathione synthetase deficiency

    MedlinePlus

    ... PubMed Njålsson R. Glutathione synthetase deficiency. Cell Mol Life Sci. 2005 Sep;62(17):1938-45. Review. Citation on PubMed Ristoff E, Larsson A. Inborn errors in the metabolism of glutathione. Orphanet J Rare Dis. 2007 Mar 30;2:16. Review. Citation on PubMed or ...

  16. Genetics Home Reference: holocarboxylase synthetase deficiency

    MedlinePlus

    ... important for the effective 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 ...

  17. Neural control of glutamine synthetase activity in rat skeletal muscles.

    PubMed

    Feng, B; Konagaya, M; Konagaya, Y; Thomas, J W; Banner, C; Mill, J; Max, S R

    1990-05-01

    The mechanism of glutamine synthetase induction in rat skeletal muscle after denervation or limb immobilization was investigated. Adult male rats were subjected to midthigh section of the sciatic nerve. At 1, 2, and 5 h and 1, 2, and 7 days after denervation, rats were killed and denervated, and contralateral control soleus and plantaris muscles were excised, weighted, homogenized, and assayed for glutamine synthetase. Glutamine synthetase activity increased approximately twofold 1 h after denervation in both muscles. By 7 days postdenervation enzyme activity had increased to three times the control level in plantaris muscle and to four times the control level in soleus muscle. Increased enzyme activity after nerve section was associated with increased maximum velocity with no change in apparent Michaelis constant. Immunotitration with an antiglutamine synthetase antibody suggested that denervation caused an increase in the number of glutamine synthetase molecules in muscle. However, Northern-blot analysis revealed no increase in the steady-state level of glutamine synthetase mRNA after denervation. A mixing experiment failed to yield evidence for the presence of a soluble factor involved in regulating the activity of glutamine synthetase in denervated muscle. A combination of denervation and dexamethasone injections resulted in additive increases in glutamine synthetase. Thus the mechanism underlying increased glutamine synthetase after denervation appears to be posttranscriptional and is distinct from that of the glucocorticoid-mediated glutamine synthetase induction previously described by us. PMID:1970709

  18. Aromatase inhibitors and anti-synthetase syndrome.

    PubMed

    Mascella, Fabio; Gianni, Lorenzo; Affatato, Alessandra; Fantini, Manuela

    2016-09-01

    Adjuvant therapy in postmenopausal women with endocrine-responsive breast cancer (BC) is actually centered on the use of anti-aromatase inhibitors (AI). Several reports, however, are emerging in literature associating the use of this drugs to rheumatic disorders. This case report describes the first case of anti-synthetase syndrome diagnosis after treatment with anti-estrogen agents in a patient with pre-existing rheumatoid arthritis. PMID:27225465

  19. 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

  20. 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.

  1. 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.

  2. Generalized orthogonal wavelet phase reconstruction.

    PubMed

    Axtell, Travis W; Cristi, Roberto

    2013-05-01

    Phase reconstruction is used for feedback control in adaptive optics systems. To achieve performance metrics for high actuator density or with limited processing capabilities on spacecraft, a wavelet signal processing technique is advantageous. Previous derivations of this technique have been limited to the Haar wavelet. This paper derives the relationship and algorithms to reconstruct phase with O(n) computational complexity for wavelets with the orthogonal property. This has additional benefits for performance with noise in the measurements. We also provide details on how to handle the boundary condition for telescope apertures. PMID:23695316

  3. 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

  4. Biosynthetic engineering of nonribosomal peptide synthetases.

    PubMed

    Kries, Hajo

    2016-09-01

    From the evolutionary melting pot of natural product synthetase genes, microorganisms elicit antibiotics, communication tools, and iron scavengers. Chemical biologists manipulate these genes to recreate similarly diverse and potent biological activities not on evolutionary time scales but within months. Enzyme engineering has progressed considerably in recent years and offers new screening, modelling, and design tools for natural product designers. Here, recent advances in enzyme engineering and their application to nonribosomal peptide synthetases are reviewed. Among the nonribosomal peptides that have been subjected to biosynthetic engineering are the antibiotics daptomycin, calcium-dependent antibiotic, and gramicidin S. With these peptides, incorporation of unnatural building blocks and modulation of bioactivities via various structural modifications have been successfully demonstrated. Natural product engineering on the biosynthetic level is not a reliable method yet. However, progress in the understanding and manipulation of biosynthetic pathways may enable the routine production of optimized peptide drugs in the near future. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. PMID:27465074

  5. 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

  6. Orthogonality catastrophe in quantum sticking.

    PubMed

    Clougherty, Dennis P; Zhang, Yanting

    2012-09-21

    We show that the orthogonality catastrophe can dramatically affect the probability with which an ultralow energy atom or ion will stick to a surface. We predict new energy-dependent scaling laws for the sticking probability in this low-energy regime. We provide numerical results of this theory for the case of ultracold electrons sticking to the surface of highly porous silicon and show that the sticking probability can differ substantially from that calculated with perturbation theory. We then generalize our results for finite surface temperatures and find surprisingly that the sticking probability can change sharply, vanishing below a critical incident energy that varies with the surface temperature. We describe in detail this superreflective surface phase for ultralow energy matter waves where the reflection coefficient is strictly equal to one. PMID:23005925

  7. Orthogonality Catastrophe in Quantum Sticking

    NASA Astrophysics Data System (ADS)

    Clougherty, Dennis P.; Zhang, Yanting

    2012-09-01

    We show that the orthogonality catastrophe can dramatically affect the probability with which an ultralow energy atom or ion will stick to a surface. We predict new energy-dependent scaling laws for the sticking probability in this low-energy regime. We provide numerical results of this theory for the case of ultracold electrons sticking to the surface of highly porous silicon and show that the sticking probability can differ substantially from that calculated with perturbation theory. We then generalize our results for finite surface temperatures and find surprisingly that the sticking probability can change sharply, vanishing below a critical incident energy that varies with the surface temperature. We describe in detail this superreflective surface phase for ultralow energy matter waves where the reflection coefficient is strictly equal to one.

  8. Comparative Biochemical and Immunological Studies of Bacterial Glutamine Synthetases

    PubMed Central

    Tronick, Steven R.; Ciardi, Joseph E.; Stadtman, E. R.

    1973-01-01

    Antisera prepared against adenylylated and unadenylylated Escherichia coli glutamine synthetase cross-reacted with the glutamine synthetases from a number of gram-negative bacteria and one gram-variable species as demonstrated by immunodiffusion and inhibition of enzyme activity. In contrast, the antisera did not cross-react with the glutamine synthetases from gram-positive bacteria (with one exception) nor with the synthetases of higher organisms. Modification of the various glutamine synthetases by covalent attachment of adenosine 5′-monophosphate (or other nucleotides) was tested for by determining whether or not snake venom phosphodiesterase altered catalytic activity in a manner similar to its effect on adenylylated E. coli glutamine synthetase. Only the activity of the glutamine synthetases from gram-negative bacteria grown with specific levels of nitrogen sources could be altered by snake venom phosphodiesterase. In addition, a relative order of antigenic homology between cross-reacting enzymes was suggested based on the patterns of spur formation in the immunodiffusion assay. Images PMID:4125585

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

    PubMed

    Schure, Mark R; Davis, Joe M

    2015-10-01

    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

  10. Energetics of S-adenosylmethionine synthetase catalysis.

    PubMed

    McQueney, M S; Anderson, K S; Markham, G D

    2000-04-18

    S-adenosylmethionine synthetase (ATP:L-methionine S-adenosyltransferase) catalyzes the only known route of biosynthesis of the primary biological alkylating agent. The internal thermodynamics of the Escherichia coli S-adenosylmethionine (AdoMet) synthetase catalyzed formation of AdoMet, pyrophosphate (PP(i)), and phosphate (P(i)) from ATP, methionine, and water have been determined by a combination of pre-steady-state kinetics, solvent isotope incorporation, and equilibrium binding measurements in conjunction with computer modeling. These studies provided the rate constants for substrate binding, the two chemical interconversion steps [AdoMet formation and subsequent tripolyphosphate (PPP(i)) hydrolysis], and product release. The data demonstrate the presence of a kinetically significant isomerization of the E.AdoMet.PP(i).P(i) complex before product release. The free energy profile for the enzyme-catalyzed reaction under physiological conditions has been constructed using these experimental values and in vivo concentrations of substrates and products. The free energy profile reveals that the AdoMet formation reaction, which has an equilibrium constant of 10(4), does not have well-balanced transition state and ground state energies. In contrast, the subsequent PPP(i) hydrolytic reaction is energetically better balanced. The thermodynamic profile indicates the use of binding energies for catalysis of AdoMet formation and the necessity for subsequent PPP(i) hydrolysis to allow enzyme turnover. Crystallographic studies have shown that a mobile protein loop gates access to the active site. The present kinetic studies indicate that this loop movement is rapid with respect to k(cat) and with respect to substrate binding at physiological concentrations. The uniformly slow binding rates of 10(4)-10(5) M(-)(1) s(-)(1) for ligands with different structures suggest that loop movement may be an intrinsic property of the protein rather than being ligand induced. PMID:10757994

  11. Molecular definition of bovine argininosuccinate synthetase deficiency.

    PubMed Central

    Dennis, J A; Healy, P J; Beaudet, A L; O'Brien, W E

    1989-01-01

    Citrullinemia is an inborn error of metabolism due to deficiency of the urea cycle enzyme, argininosuccinate synthetase [L-citrulline:L-aspartate ligase (AMP-forming), EC 6.3.4.5]. The disease was first described in humans but was recently reported in dairy cattle in Australia. Here we report the nucleotide sequence of the normal bovine cDNA for argininosuccinate synthetase and the mutation present in animals with citrullinemia. Analysis of DNA from affected animals by Southern blotting did not readily identify the mutation in the bovine gene. RNA (Northern) blotting revealed a major reduction in the steady-state amount of mRNA in the liver of affected animals to less than 5% of controls. The bovine cDNA was cloned and sequenced and revealed 96% identity with the deduced human sequence at the amino acid level. Starting with mutant bovine liver, the mRNA was reverse-transcribed; the cDNA product was amplified with the polymerase chain reaction, cloned, and sequenced. The sequence revealed a C----T transition converting arginine-86 (CGA) to a nonsense codon (TGA). A second C----T transition represented a polymorphism in proline-175 (CCC----CCT). The mutation and the polymorphism were confirmed by amplification of genomic DNA and demonstration with restriction endonuclease enzymes of both the loss of an Ava II site in DNA from mutant animals at codon 86 and the presence or absence of a Dde I site at codon 175. The loss of the Ava II site can be used for rapid, economical, nonradioactive detection of heterozygotes for bovine citrullinemia. Images PMID:2813370

  12. Genetics Home Reference: carbamoyl phosphate synthetase I deficiency

    MedlinePlus

    Skip to main content Your Guide to Understanding Genetic Conditions Enable Javascript for addthis links to activate. ... Conditions Genes Chromosomes & mtDNA Resources Help Me Understand Genetics Home Health Conditions carbamoyl phosphate synthetase I deficiency ...

  13. Continuous spectrophotometric assay for aminoacyl-tRNA synthetases.

    PubMed

    Chang, G G; Pan, F; Lin, Y H; Wang, H Y

    1984-11-01

    A simple, continuous assay for aminoacyl-tRNA synthetases utilizing a commercially available pyrophosphate assay reagent kit was demonstrated. The method coupled aminoacyl-tRNA synthetase activity with pyrophosphate-dependent fructose-6-phosphate kinase, aldolase, triosephosphate isomerase, and glycerophosphate dehydrogenase. PPi formation was correlated with the oxidation of NADH, and was monitored continuously by the decrease of absorbance at 340 nm. PMID:6099060

  14. Organisation and sequence determination of glutamine-dependent carbamoyl phosphate synthetase II in Toxoplasma gondii.

    PubMed

    Fox, Barbara A; Bzik, David J

    2003-01-01

    Carbamoyl phosphate synthetase II encodes the first enzymic step of de novo pyrimidine biosynthesis. Carbamoyl phosphate synthetase II is essential for Toxoplasma gondii replication and virulence. In this study, we characterised the primary structure of a 28kb gene encoding Toxoplasma gondii carbamoyl phosphate synthetase II. The carbamoyl phosphate synthetase II gene was interrupted by 36 introns. The predicted protein encoded by the 37 carbamoyl phosphate synthetase II exons was a 1,687 amino acid polypeptide with an N-terminal glutamine amidotransferase domain fused with C-terminal carbamoyl phosphate synthetase domains. This bifunctional organisation of carbamoyl phosphate synthetase II is unique, so far, to protozoan parasites from the phylum Apicomplexa (Plasmodium, Babesia, Toxoplasma) or zoomastigina (Trypanosoma, Leishmania). Apicomplexan parasites possessed the largest carbamoyl phosphate synthetase II enzymes due to insertions in the glutamine amidotransferase and carbamoyl phosphate synthetase domains that were not present in the corresponding gene segments from bacteria, plants, fungi and mammals. The C-terminal allosteric regulatory domain, the carbamoyl phosphate synthetase linker domain and the oligomerisation domain were also distinct from the corresponding domains in other species. The novel C-terminal regulatory domain may explain the lack of activation of Toxoplasma gondii carbamoyl phosphate synthetase II by the allosteric effector 5-phosphoribosyl 1-pyrophosphate. Toxoplasma gondii growth in vitro was markedly inhibited by the glutamine antagonist acivicin, an inhibitor of glutamine amidotransferase activity typically associated with carbamoyl phosphate synthetase II, guanosine monophosphate synthetase, or CTP synthetase. PMID:12547350

  15. [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. PMID:26964231

  16. 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.

  17. 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.

  18. Kinetics profiling of gramicidin S synthetase A, a member of nonribosomal peptide synthetases.

    PubMed

    Sun, Xun; Li, Hao; Alfermann, Jonas; Mootz, Henning D; Yang, Haw

    2014-12-23

    Nonribosomal peptide synthetases (NRPS) incorporate assorted amino acid substrates into complex natural products. The substrate is activated via the formation of a reactive aminoacyl adenylate and is subsequently attached to the protein template via a thioester bond. The reactive nature of such intermediates, however, leads to side reactions that also break down the high-energy anhydride bond. The off-pathway kinetics or their relative weights compared to that of the on-pathway counterpart remains generally elusive. Here, we introduce multiplatform kinetics profiling to quantify the relative weights of on- and off-pathway reactions. Using the well-defined stoichiometry of thioester formation, we integrate a mass spectrometry (MS) kinetics assay, a high-performance liquid chromatography (HPLC) assay, and an ATP-pyrophosphate (PPi) exchange assay to map out a highly efficient on-pathway kinetics profile of the substrate activation and intermediate uploading (>98% relative weight) for wide-type gramicidin S synthetase A (GrsA) and a 87% rate profile for a cysteine-free GrsA mutant. Our kinetics profiling approach complements the existing enzyme-coupled byproduct-release assays, unraveling new mechanistic insights of substrate activation/channeling in NRPS enzymes. PMID:25437123

  19. 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.

  20. Regulation of Glutamine Synthetase V. Partial Purification and Properties of Glutamine Synthetase from Bacillus licheniformis

    PubMed Central

    Hubbard, Jerry S.; Stadtman, E. R.

    1967-01-01

    The glutamine synthetase of Bacillus licheniformis has been obtained at about 15% purity. Sucrose gradient centrifugation gave a molecular weight value of approximately 612,000. Both l- and d-glutamate can be utilized as substrates in the biosynthetic reaction, although the l isomer was five times more active. The requirement for adenosine triphosphate (ATP) can be partially replaced by guanosine or inosine triphosphates, but not by cytidine or uridine triphosphates. The Mn++ was required for activity, and the requirement cannot be satisfied with Mg++. Maximal activity of the biosynthetic reaction was observed when ATP and Mn++ were present in equimolar amounts. An excess of either reactant gave less activity. However, other purine and pyrimidine nucleotides, when added in combination with ATP, can partially substitute for ATP in attaining the equimolar ratio of nucleotide to Mn++. A complex of ATP and Mn++ is the preferred form of substrate. The B. licheniformis enzyme catalyzes the glutamyl transfer reaction but at a much slower rate than the Escherichia coli glutamine synthetase. Either adenosine diphosphate (ADP) or ATP can activate the glutamotransferase, although ADP is more active. PMID:6051339

  1. Orthogonality and distinguishability: Criterion for local distinguishability of arbitrary orthogonal states

    SciTech Connect

    Chen Pingxing; Li Chengzu

    2003-12-01

    We consider the relation between the orthogonality and the distinguishability of a set of arbitrary states (including multipartite states). It is shown that if a set of arbitrary states can be distinguished by local operations and classical communication (LOCC), each of the states can be written as a linear combination of product vectors such that all product vectors of one of the states are orthogonal to the other states. With this result we then prove a simple necessary condition for LOCC distinguishability of a class of orthogonal states. These conclusions may be useful in discussing the distinguishability of orthogonal quantum states further, understanding the essence of nonlocality and discussing the distillation of entanglement.

  2. 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

  3. [Thromboxane A2 synthetase inhibitor in asthma therapy].

    PubMed

    Machida, K; Takagi, K; Horiba, M

    1996-11-01

    Thromboxane A2(TXA2), a platelet aggregator and vasoconstricter, has been implicated as a potential mediator of bronchial asthma. TXA2 induces potent contraction of airway smooth muscles and airway hyperresponsiveness. OKY-046 (ozagrel hydrochloride) is a specific inhibitor of TXA2 synthetase and a new antiasthmatic agent. In a phase III study ozagrel has shown significantly higher effect in ameliorating the asthma symptoms and reduced the dose of concomitant steroid therapy compared to azelastine hydrochloride. Both basical and clinical studies showed that TXA2 synthetase inhibitor is effective on airway hyperresponsiveness. In this review the role of TXA2 synthetase inhibitor in current asthma therapy, which is based on the Japanese guideline of allergic disorders, was discussed. PMID:8950950

  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. 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.

  6. 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. PMID:27581854

  7. Kyotorphin (tyrosine-arginine) synthetase in rat brain synaptosomes.

    PubMed

    Ueda, H; Yoshihara, Y; Fukushima, N; Shiomi, H; Nakamura, A; Takagi, H

    1987-06-15

    Kyotorphin (Tyr-Arg) is a unique neuropeptide which produces analgesia by releasing Met-enkephalin from slices of the brain and spinal cord. Recent studies revealed that kyotorphin possesses the properties of neurotransmitter/neuroregulator. In the present study, we identified a kyotorphin synthetase in the soluble fraction of rat brain synaptosomes (synaptosol) and characterized it. The enzyme partially purified with Sephacryl S-300 showed an absolute requirement for ATP, MgCl2, tyrosine, and arginine. The optimal pH was 7.5-9.0 and the pI was determined to be 6.1-6.2 by isoelectric focusing. The Km was 25.6 microM for tyrosine, 926 microM for arginine, 294 microM for ATP, and 442 microM for MgCl2. The Vmax was 34.0 pmol/mg of protein/h. The apparent molecular size of this "kyotorphin synthetase" further purified by the DE52 column was 240,000-245,000 daltons, estimated using TSKgel G4000SW column chromatography. The enzyme reaction is represented by the following equation: Tyr + Arg + ATP + MgCl2 + kyotorphin synthetase----Tyr-Arg (kyotorphin) + AMP + PPi + MgCl2 + kyotorphin synthetase. The regional distribution and subcellular localization of the synthetase showed a close correlation to that of kyotorphin levels in the rat brain. The amounts of kyotorphin formed from amino acids by the synthetase in the dialyzed synaptosol was 3.0-4.0 times higher than that from precursor proteins by processing enzymes within the 30 min incubation. PMID:3597366

  8. Nonlocality of orthogonal product basis quantum states

    NASA Astrophysics Data System (ADS)

    Zhang, Zhi-Chao; Gao, Fei; Tian, Guo-Jing; Cao, Tian-Qing; Wen, Qiao-Yan

    2014-08-01

    In this paper, we mainly study the local indistinguishability of mutually orthogonal product basis quantum states in the high-dimensional quantum systems. In the Hilbert space of 3⊗3, Walgate and Hardy [Phys. Rev. Lett. 89, 147901 (2002), 10.1103/PhysRevLett.89.147901] presented a very simple proof for nonlocality of nine orthogonal product basis quantum states which are given by Bennett et al. [Phys. Rev. A 59, 1070 (1999), 10.1103/PhysRevA.59.1070]. In the quantum system of d⊗d, where d is odd, we construct d2 orthogonal product basis quantum states and prove these states are locally indistinguishable. Then we are able to construct some locally indistinguishable product basis quantum states in the multipartite systems. All these results reveal the phenomenon of "nonlocality without entanglement."

  9. Orthogonal rational functions and tridiagonal matrices

    NASA Astrophysics Data System (ADS)

    Bultheel, A.; González-Vera, P.; Hendriksen, E.; Njåstad, O.

    2003-04-01

    We study the recurrence relation for rational functions whose poles are in a prescribed sequence of numbers that are real or infinite and that are orthogonal with respect to an Hermitian positive linear functional. We especially discuss the interplay between finite and infinite poles. The recurrence relation will also be described in terms of a tridiagonal matrix which is a generalization of the Jacobi. matrix of the polynomial situation which corresponds to placing all the poles at infinity. This matrix not only describes the recurrence relation, but it can be used to give a determinant expression for the orthogonal rational functions and it also allows for the formulation of a generalized eigenvalue problem whose eigenvalues are the zeros of an orthogonal rational function. These nodes can be used in rational Gauss-type quadrature formulas and the corresponding weights can be obtained from the first components of the corresponding eigenvectors.

  10. Improved piecewise orthogonal signal correction algorithm.

    PubMed

    Feudale, Robert N; Tan, Huwei; Brown, Steven D

    2003-10-01

    Piecewise orthogonal signal correction (POSC), an algorithm that performs local orthogonal filtering, was recently developed to process spectral signals. POSC was shown to improve partial leastsquares regression models over models built with conventional OSC. However, rank deficiencies within the POSC algorithm lead to artifacts in the filtered spectra when removing two or more POSC components. Thus, an updated OSC algorithm for use with the piecewise procedure is reported. It will be demonstrated how the mathematics of this updated OSC algorithm were derived from the previous version and why some OSC versions may not be as appropriate to use with the piecewise modeling procedure as the algorithm reported here. PMID:14639746

  11. Phosphorylation of five aminoacyl-tRNA synthetases in reticulocytes and identification of the protein kinases phosphorylating threonyl-tRNA synthetase from rat liver

    SciTech Connect

    Pendergast, A.M.; Traugh, J.A.

    1986-05-01

    Five aminoacyl-tRNA synthetases in the high molecular weight complex were phosphorylated in rabbit reticulocytes following labeling with /sup 32/P. The five synthetases phosphorylated were the glutamyl-, glutaminyl-, lysyl-, aspartyl- and methionyl-tRNA synthetases. In addition, a 37,000 dalton protein, associated with the synthetase complex and tentatively identified as casein kinase I, was also phosphorylated in intact cells. Phosphoamino acid analysis of the proteins indicated all of the phosphate was on seryl residues. Incubation of reticulocytes with /sup 32/P in the presence of 8-bromo-cAMP and o, the 3-isobutyl-1-methylxanthine resulted in a six-fold increase in phosphorylation of the glutaminyl-tRNA synthetase, a two-fold increase in phosphorylation of the aspartyl-tRNA synthetase, and a 50 to 60% decrease in phosphorylation of the glutamyl-, methionyl- and lysyl-tRNA synthetases and the M/sub r/ 37,000 protein. When the site(s) on the glutaminyl-tRNA synthetase phosphorylated in response to 8-bromo-cAMP was analyzed by two-dimensional tryptic phosphopeptide mapping, a single phosphopeptide was observed which was identical to that obtained in vitro upon phosphorylation with the cAMP-dependent protein kinase. Also, the authors identify here, the protein kinases phosphorylating threonyl-tRNA synthetase from rat liver. They are protease activated kinase I, the cAMP-dependent protein kinase and protein kinase C.

  12. 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

  13. 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. PMID:25385624

  14. Genetic and Immunological Studies of Bacteriophage T4 Thymidylate Synthetase

    PubMed Central

    Krauss, S. W.; Stollar, B. D.; Friedkin, M.

    1973-01-01

    Thymidylate synthetase, which appears after infection of Escherichia coli with bacteriophage T4, has been partially purified. The phage enzyme is immunologically distinct from the host enzyme and has a molecular weight of 50,000 in comparison to 68,000 for the host enzyme. A system has been developed to characterize T4 td mutants previously known to have impaired expression of phage thymidylate synthetase. For this system, an E. coli host lacking thymidylate synthetase was isolated. Known genetic suppressors were transduced into this host. The resulting isogenic hosts were infected with phage T4 td mutants. The specific activities and amounts of cross-reacting material induced by several different types of phage mutants under conditions of suppression or non-suppression have been examined. The results show that the phage carries the structural gene specifying the thymidylate synthetase which appears after phage infection, and that the combination of plaque morphology, enzyme activity assays, and an assay for immunologically cross-reacting material provides a means for identifying true amber mutants of the phage gene. Images PMID:4575286

  15. N-acetylaspartylglutamate synthetase II synthesizes N-acetylaspartylglutamylglutamate.

    PubMed

    Lodder-Gadaczek, Julia; Becker, Ivonne; Gieselmann, Volkmar; Wang-Eckhardt, Lihua; Eckhardt, Matthias

    2011-05-13

    N-Acetylaspartylglutamate (NAAG) is found at high concentrations in the vertebrate nervous system. NAAG is an agonist at group II metabotropic glutamate receptors. In addition to its role as a neuropeptide, a number of functions have been proposed for NAAG, including a role as a non-excitotoxic transport form of glutamate and a molecular water pump. We recently identified a NAAG synthetase (now renamed NAAG synthetase I, NAAGS-I), encoded by the ribosomal modification protein rimK-like family member B (Rimklb) gene, as a member of the ATP-grasp protein family. We show here that a structurally related protein, encoded by the ribosomal modification protein rimK-like family member A (Rimkla) gene, is another NAAG synthetase (NAAGS-II), which in addition, synthesizes the N-acetylated tripeptide N-acetylaspartylglutamylglutamate (NAAG(2)). In contrast, NAAG(2) synthetase activity was undetectable in cells expressing NAAGS-I. Furthermore, we demonstrate by mass spectrometry the presence of NAAG(2) in murine brain tissue and sciatic nerves. The highest concentrations of both, NAAG(2) and NAAG, were found in sciatic nerves, spinal cord, and the brain stem, in accordance with the expression level of NAAGS-II. To our knowledge the presence of NAAG(2) in the vertebrate nervous system has not been described before. The physiological role of NAAG(2), e.g. whether it acts as a neurotransmitter, remains to be determined. PMID:21454531

  16. 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.

  17. Nonlinear Submodels Of Orthogonal Linear Models

    ERIC Educational Resources Information Center

    Bechtel, Gordon G.

    1973-01-01

    It is the purpose of this paper to suggest the orthogonal analysis of variance as a device for simplifying either the analytic or iterative problem of finding LS (least squares) estimates for the parameters of particular nonlinear models. (Author/RK)

  18. 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.

  19. The Rigid Orthogonal Procrustes Rotation Problem

    ERIC Educational Resources Information Center

    ten Berge, Jos M. F.

    2006-01-01

    The problem of rotating a matrix orthogonally to a best least squares fit with another matrix of the same order has a closed-form solution based on a singular value decomposition. The optimal rotation matrix is not necessarily rigid, but may also involve a reflection. In some applications, only rigid rotations are permitted. Gower (1976) has…

  20. Border separation for adjacent orthogonal fields

    SciTech Connect

    Werner, B.L.; Khan, F.M.; Sharma, S.C.; Lee, C.K.; Kim, T.H. )

    1991-06-01

    Field border separations for adjacent orthogonal fields can be calculated geometrically, given the validity of some important assumptions such as beam alignment and field uniformity. Thermoluminescent dosimetry (TLD) measurements were used to investigate dose uniformity across field junctions as a function of field separation and, in particular, to review the CCSG recommendation for the treatment of medulloblastoma with separate head and spine fields.

  1. Three-Dimensional Orthogonal Co-ordinates

    ERIC Educational Resources Information Center

    Astin, J.

    1974-01-01

    A systematic approach to general orthogonal co-ordinates, suitable for use near the end of a beginning vector analysis course, is presented. It introduces students to tensor quantities and shows how equations and quantities needed in classical problems can be determined. (Author/LS)

  2. A class of orthogonal nonrecursive binomial filters.

    NASA Technical Reports Server (NTRS)

    Haddad, R. A.

    1971-01-01

    The time- and frequency-domain properties of the orthogonal binomial sequences are presented. It is shown that these sequences, or digital filters based on them, can be generated using adders and delay elements only. The frequency-domain behavior of these nonrecursive binomial filters suggests a number of applications as low-pass Gaussian filters or as inexpensive bandpass filters.

  3. 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)

  4. Orthogonal Thin Film Photovoltaics on Vertical Nanostructures.

    PubMed

    Ahnood, Arman; Zhou, H; Suzuki, Y; Sliz, R; Fabritius, T; Nathan, Arokia; Amaratunga, G A J

    2015-12-01

    Decoupling paths of carrier collection and illumination within photovoltaic devices is one promising approach for improving their efficiency by simultaneously increasing light absorption and carrier collection efficiency. Orthogonal photovoltaic devices are core-shell type structures consisting of thin film photovoltaic stack on vertical nanopillar scaffolds. These types of devices allow charge collection to take place in the radial direction, perpendicular to the path of light in the vertical direction. This approach addresses the inherently high recombination rate of disordered thin films, by allowing semiconductor films with minimal thicknesses to be used in photovoltaic devices, without performance degradation associated with incomplete light absorption. This work considers effects which influence the performance of orthogonal photovoltaic devices. Illumination non-uniformity as light travels across the depth of the pillars, electric field enhancement due to the nanoscale size and shape of the pillars, and series resistance due to the additional surface structure created through the use of pillars are considered. All of these effects influence the operation of orthogonal solar cells and should be considered in the design of vertically nanostructured orthogonal photovoltaics. PMID:26676997

  5. Orthogonal Thin Film Photovoltaics on Vertical Nanostructures

    NASA Astrophysics Data System (ADS)

    Ahnood, Arman; Zhou, H.; Suzuki, Y.; Sliz, R.; Fabritius, T.; Nathan, Arokia; Amaratunga, G. A. J.

    2015-12-01

    Decoupling paths of carrier collection and illumination within photovoltaic devices is one promising approach for improving their efficiency by simultaneously increasing light absorption and carrier collection efficiency. Orthogonal photovoltaic devices are core-shell type structures consisting of thin film photovoltaic stack on vertical nanopillar scaffolds. These types of devices allow charge collection to take place in the radial direction, perpendicular to the path of light in the vertical direction. This approach addresses the inherently high recombination rate of disordered thin films, by allowing semiconductor films with minimal thicknesses to be used in photovoltaic devices, without performance degradation associated with incomplete light absorption. This work considers effects which influence the performance of orthogonal photovoltaic devices. Illumination non-uniformity as light travels across the depth of the pillars, electric field enhancement due to the nanoscale size and shape of the pillars, and series resistance due to the additional surface structure created through the use of pillars are considered. All of these effects influence the operation of orthogonal solar cells and should be considered in the design of vertically nanostructured orthogonal photovoltaics.

  6. Lincosamide synthetase--a unique condensation system combining elements of nonribosomal peptide synthetase and mycothiol metabolism.

    PubMed

    Janata, Jiri; Kadlcik, Stanislav; Koberska, Marketa; Ulanova, Dana; Kamenik, Zdenek; Novak, Petr; Kopecky, Jan; Novotna, Jitka; Radojevic, Bojana; Plhackova, Kamila; Gazak, Radek; Najmanova, Lucie

    2015-01-01

    In the biosynthesis of lincosamide antibiotics lincomycin and celesticetin, the amino acid and amino sugar units are linked by an amide bond. The respective condensing enzyme lincosamide synthetase (LS) is expected to be an unusual system combining nonribosomal peptide synthetase (NRPS) components with so far unknown amino sugar related activities. The biosynthetic gene cluster of celesticetin was sequenced and compared to the lincomycin one revealing putative LS coding ORFs shared in both clusters. Based on a bioassay and production profiles of S. lincolnensis strains with individually deleted putative LS coding genes, the proteins LmbC, D, E, F and V were assigned to LS function. Moreover, the newly recognized N-terminal domain of LmbN (LmbN-CP) was also assigned to LS as a NRPS carrier protein (CP). Surprisingly, the homologous CP coding sequence in celesticetin cluster is part of ccbZ gene adjacent to ccbN, the counterpart of lmbN, suggesting the gene rearrangement, evident also from still active internal translation start in lmbN, and indicating the direction of lincosamide biosynthesis evolution. The in vitro test with LmbN-CP, LmbC and the newly identified S. lincolnensis phosphopantetheinyl transferase Slp, confirmed the cooperation of the previously characterized NRPS A-domain LmbC with a holo-LmbN-CP in activation of a 4-propyl-L-proline precursor of lincomycin. This result completed the functional characterization of LS subunits resembling NRPS initiation module. Two of the four remaining putative LS subunits, LmbE/CcbE and LmbV/CcbV, exhibit low but significant homology to enzymes from the metabolism of mycothiol, the NRPS-independent system processing the amino sugar and amino acid units. The functions of particular LS subunits as well as cooperation of both NRPS-based and NRPS-independent LS blocks are discussed. The described condensing enzyme represents a unique hybrid system with overall composition quite dissimilar to any other known enzyme system

  7. 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.

  8. Impact of radar systematic error on the orthogonal frequency division multiplexing chirp waveform orthogonality

    NASA Astrophysics Data System (ADS)

    Wang, Jie; Liang, Xingdong; Chen, Longyong; Ding, Chibiao

    2015-01-01

    Orthogonal frequency division multiplexing (OFDM) chirp waveform, which is composed of two successive identical linear frequency modulated subpulses, is a newly proposed orthogonal waveform scheme for multiinput multioutput synthetic aperture radar (SAR) systems. However, according to the waveform model, radar systematic error, which introduces phase or amplitude difference between the subpulses of the OFDM waveform, significantly degrades the orthogonality. The impact of radar systematic error on the waveform orthogonality is mainly caused by the systematic nonlinearity rather than the thermal noise or the frequency-dependent systematic error. Due to the influence of the causal filters, the first subpulse leaks into the second one. The leaked signal interacts with the second subpulse in the nonlinear components of the transmitter. This interaction renders a dramatic phase distortion in the beginning of the second subpulse. The resultant distortion, which leads to a phase difference between the subpulses, seriously damages the waveform's orthogonality. The impact of radar systematic error on the waveform orthogonality is addressed. Moreover, the impact of the systematic nonlinearity on the waveform is avoided by adding a standby between the subpulses. Theoretical analysis is validated by practical experiments based on a C-band SAR system.

  9. 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

  10. 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

  11. Describing freeform surfaces with orthogonal functions

    NASA Astrophysics Data System (ADS)

    Ochse, D.; Uhlendorf, K.; Reichmann, L.

    2015-09-01

    In optical design with freeform surfaces descriptions of the surfaces are needed that use only few parameters and are suitable for optimisation. Depending on the merit function - spot size or wavefront error - and the position of the surface in the system, different surface types can yield different optimisation performance. It has been demonstrated by G. Forbes that slope orthogonal polynomials are an advantageous freeform description. From literature on Gaussian moments it is known that this can be achieved using differences of Zernike polynomials, which are easy to compute and implement with recent algorithms. We will demonstrate the benefits of Zernike polynomials with optimisation examples. Furthermore we present an orthogonal surface representation on a rectangular aperture based on Chebyshev polynomials. This description is very convenient when the aperture has a very high aspect ratio, or when designing a system with a rectangular pupil.

  12. 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. PMID:26390483

  13. Orthogonal gradient networks via post polymerization reaction

    NASA Astrophysics Data System (ADS)

    Chinnayan Kannan, Pandiyarajan; Genzer, Jan

    2015-03-01

    We report a novel synthetic route to generate orthogonal gradient networks through post polymerization reaction using pentaflurophenylmethacrylate (PFPMAc) active ester chemistry. These chemoselective monomers were successfully copolymerized with 5 mole% of the photo (methacryloyloxybenzophenone) and thermal (styrenesulfonylazide) crosslinkers. Subsequently, the copolymers were modified by a series of amines having various alkyl chain lengths. The conversion of post polymerization reaction was monitored using Fourier Transform Infrared Spectroscopy (FT-IR) and noticed that almost all pentaflurophenyl moieties are substituted by amines within in an hour without affecting the crosslinkers. In addition, the incorporation of photo and thermal crosslinkers in the polymer enabled us to achieve stable and covalently surface-bound polymer gradient networks (PGN) in an orthogonal manner, i.e. complete control over the crosslink density of the network in two opposite directions (i.e. heat vs photo). The network properties such as wettability, swelling and tensile modulus of the gradient coatings are studied and revealed in the paper.

  14. Orthogonalized operators for the f shell

    SciTech Connect

    Judd, B.R.; Crosswhite, H.

    1984-04-01

    Orthogonalized operators are introduced in the atomic configurations f/sup N/ in order to yield parameters that are more precisely defined and more stable than the conventional ones. Of the four Racah operators e/sub 0/, e/sub 1/, e/sub 2/, only e/sub 1/ needs adjusting. The set of two-electron scalars is made complete by the generalized Trees operators e/sub ..cap alpha../', e/sub ..beta../', and e/sub ..gamma../'. Of the three-electron scalars t/sub i/, only t/sub 2/ requires alteration. The theory is illustrated for f/sup 3/ by adding the orthogonalized operators in successive steps and comparing the fits with those obtained if the conventional operators are used.

  15. Orthogonal nilpotent superfields from linear models

    NASA Astrophysics Data System (ADS)

    Kallosh, Renata; Karlsson, Anna; Mosk, Benjamin; Murli, Divyanshu

    2016-05-01

    We derive supersymmetry/supergravity models with constrained orthogonal nilpotent superfields from the linear models in the formal limit where the masses of the sgoldstino, inflatino and sinflaton tend to infinity. The case where the sinflaton mass remains finite leads to a model with a `relaxed' constraint, where the sinflaton remains an independent field. Our procedure is equivalent to a requirement that some of the components of the curvature of the moduli space tend to infinity.

  16. Stochastic processes with orthogonal polynomial eigenfunctions

    NASA Astrophysics Data System (ADS)

    Griffiths, Bob

    2009-12-01

    Markov processes which are reversible with either Gamma, Normal, Poisson or Negative Binomial stationary distributions in the Meixner class and have orthogonal polynomial eigenfunctions are characterized as being processes subordinated to well-known diffusion processes for the Gamma and Normal, and birth and death processes for the Poisson and Negative Binomial. A characterization of Markov processes with Beta stationary distributions and Jacobi polynomial eigenvalues is also discussed.

  17. Orthogonal rational functions and quadrature on an interval

    NASA Astrophysics Data System (ADS)

    van Deun, J.; Bultheel, A.

    2003-04-01

    Rational functions with real poles and poles in the complex lower half-plane, orthogonal on the real line, are well known. Quadrature formulas similar to the Gauss formulas for orthogonal polynomials have been studied. We generalize to the case of arbitrary complex poles and study orthogonality on a finite interval. The zeros of the orthogonal rational functions are shown to satisfy a quadratic eigenvalue problem. In the case of real poles, these zeros are used as nodes in the quadrature formulas.

  18. 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.

  19. Nested Krylov methods and preserving the orthogonality

    NASA Technical Reports Server (NTRS)

    Desturler, Eric; Fokkema, Diederik R.

    1993-01-01

    Recently the GMRESR inner-outer iteraction scheme for the solution of linear systems of equations was proposed by Van der Vorst and Vuik. Similar methods have been proposed by Axelsson and Vassilevski and Saad (FGMRES). The outer iteration is GCR, which minimizes the residual over a given set of direction vectors. The inner iteration is GMRES, which at each step computes a new direction vector by approximately solving the residual equation. However, the optimality of the approximation over the space of outer search directions is ignored in the inner GMRES iteration. This leads to suboptimal corrections to the solution in the outer iteration, as components of the outer iteration directions may reenter in the inner iteration process. Therefore we propose to preserve the orthogonality relations of GCR in the inner GMRES iteration. This gives optimal corrections; however, it involves working with a singular, non-symmetric operator. We will discuss some important properties, and we will show by experiments that, in terms of matrix vector products, this modification (almost) always leads to better convergence. However, because we do more orthogonalizations, it does not always give an improved performance in CPU-time. Furthermore, we will discuss efficient implementations as well as the truncation possibilities of the outer GCR process. The experimental results indicate that for such methods it is advantageous to preserve the orthogonality in the inner iteration. Of course we can also use iteration schemes other than GMRES as the inner method; methods with short recurrences like GICGSTAB are of interest.

  20. Functional systems with orthogonal dynamic covalent bonds.

    PubMed

    Wilson, Adam; Gasparini, Giulio; Matile, Stefan

    2014-03-21

    This review summarizes the use of orthogonal dynamic covalent bonds to build functional systems. Dynamic covalent bonds are unique because of their dual nature. They can be as labile as non-covalent interactions or as permanent as covalent bonds, depending on conditions. Examples from nature, reaching from the role of disulfides in protein folding to thioester exchange in polyketide biosynthesis, indicate how dynamic covalent bonds are best used in functional systems. Several synthetic functional systems that employ a single type of dynamic covalent bonds have been reported. Considering that most functional systems make simultaneous use of several types of non-covalent interactions together, one would expect the literature to contain many examples in which different types of dynamic covalent bonds are similarly used in tandem. However, the incorporation of orthogonal dynamic covalent bonds into functional systems is a surprisingly rare and recent development. This review summarizes the available material comprehensively, covering a remarkably diverse collection of functions. However, probably more revealing than the specific functions addressed is that the questions asked are consistently quite unusual, very demanding and highly original, focusing on molecular systems that can self-sort, self-heal, adapt, exchange, replicate, transcribe, or even walk and "think" (logic gates). This focus on adventurous chemistry off the beaten track supports the promise that with orthogonal dynamic covalent bonds we can ask questions that otherwise cannot be asked. The broad range of functions and concepts covered should appeal to the supramolecular organic chemist but also to the broader community. PMID:24287608

  1. 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.

  2. Orthogonal NGS for High Throughput Clinical Diagnostics

    PubMed Central

    Chennagiri, Niru; White, Eric J.; Frieden, Alexander; Lopez, Edgardo; Lieber, Daniel S.; Nikiforov, Anastasia; Ross, Tristen; Batorsky, Rebecca; Hansen, Sherry; Lip, Va; Luquette, Lovelace J.; Mauceli, Evan; Margulies, David; Milos, Patrice M.; Napolitano, Nichole; Nizzari, Marcia M.; Yu, Timothy; Thompson, John F.

    2016-01-01

    Next generation sequencing is a transformative technology for discovering and diagnosing genetic disorders. However, high-throughput sequencing remains error-prone, necessitating variant confirmation in order to meet the exacting demands of clinical diagnostic sequencing. To address this, we devised an orthogonal, dual platform approach employing complementary target capture and sequencing chemistries to improve speed and accuracy of variant calls at a genomic scale. We combined DNA selection by bait-based hybridization followed by Illumina NextSeq reversible terminator sequencing with DNA selection by amplification followed by Ion Proton semiconductor sequencing. This approach yields genomic scale orthogonal confirmation of ~95% of exome variants. Overall variant sensitivity improves as each method covers thousands of coding exons missed by the other. We conclude that orthogonal NGS offers improvements in variant calling sensitivity when two platforms are used, better specificity for variants identified on both platforms, and greatly reduces the time and expense of Sanger follow-up, thus enabling physicians to act on genomic results more quickly. PMID:27090146

  3. Constrained predictive control using orthogonal expansions

    SciTech Connect

    Finn, C.K. ); Wahlberg, B. . Dept. of Automatic Control); Ydstie, B.E. . Dept. of Chemical Engineering)

    1993-11-01

    Orthogonal expansion is routinely used for multivariable predictive control and optimization in the chemical and petrochemical manufacturing industries. In this article, the authors approximate bounded operators by orthogonal expansion. The rate of convergence depends on the choice of basis functions. Markov-Laguerre functions give rapid convergence for open-loop stable systems with long delay. The Markov-Kautz model can be used for lightly damped systems, and a more general orthogonal expansion is developed for modeling multivariable systems with widely scattered poles. The finite impulse response model is a special case of these models. A-priori knowledge about dominant time constants, time delay and oscillatory modes is used to reduce the model complexity and to improve conditioning of the parameter estimation algorithm. Algorithms for predictive control are developed, as well as conditions for constraint compatibility, closed-loop stability and constraint satisfaction for the ideal case. An H[infinity]--like design technique proposed guarantees robust stability in the presence of input constraints; output constraints may give chatter. A chatter-free algorithm is proposed.

  4. Glutathione production by recombinant Escherichia coli expressing bifunctional glutathione synthetase.

    PubMed

    Wang, Dezheng; Wang, Cheng; Wu, Hui; Li, Zhimin; Ye, Qin

    2016-01-01

    Glutathione (GSH) is an important bioactive substance applied widely in pharmaceutical and food industries. Due to the strong product inhibition in the GSH biosynthetic pathway, high levels of intracellular content, yield and productivity of GSH are difficult to achieve. Recently, a novel bifunctional GSH synthetase was identified to be less sensitive to GSH. A recombinant Escherichia coli strain expressing gshF encoding the bifunctional glutathione synthetase of Streptococcus thermophilus was constructed for GSH production. In this study, efficient GSH production using this engineered strain was investigated. The cultivation process was optimized by controlling dissolved oxygen (DO), amino acid addition and glucose feeding. 36.8 mM (11.3 g/L) GSH were formed at a productivity of 2.06 mM/h when the amino acid precursors (75 mM each) were added and glucose was supplied as the sole carbon and energy source. PMID:26586402

  5. Glutamine synthetase gene evolution: a good molecular clock.

    PubMed Central

    Pesole, G; Bozzetti, M P; Lanave, C; Preparata, G; Saccone, C

    1991-01-01

    Glutamine synthetase (EC 6.3.1.2) gene evolution in various animals, plants, and bacteria was evaluated by a general stationary Markov model. The evolutionary process proved to be unexpectedly regular even for a time span as long as that between the divergence of prokaryotes from eukaryotes. This enabled us to draw phylogenetic trees for species whose phylogeny cannot be easily reconstructed from the fossil record. Our calculation of the times of divergence of the various organelle-specific enzymes led us to hypothesize that the pea and bean chloroplast genes for these enzymes originated from the duplication of nuclear genes as a result of the different metabolic needs of the various species. Our data indicate that the duplication of plastid glutamine synthetase genes occurred long after the endosymbiotic events that produced the organelles themselves. PMID:1671172

  6. 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

  7. Glutamine synthetase of Klebsiella aerogenes: properties of glnD mutants lacking uridylyltransferase.

    PubMed Central

    Foor, F; Cedergren, R J; Streicher, S L; Rhee, S G; Magasanik, B

    1978-01-01

    The glnD mutation of Klebsiella aerogenes is cotransducible by phage P1 with pan (requirement for pantothenate) and leads to a loss of uridylytransferase and uridylyl-removing enzyme, components of the glutamine synthetase adenylylation system. This defect results in an inability to deadenylylate glutamine synthetase rapidly and in a requirement for glutamine for normal growth. Suppression of the glnD mutation are located at the glutamine synthetase structural gene glnA. PMID:26659

  8. 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.

  9. Cysteinyl-tRNA synthetase: determination of the last E. coli aminoacyl-tRNA synthetase primary structure.

    PubMed Central

    Eriani, G; Dirheimer, G; Gangloff, J

    1991-01-01

    The gene coding for E. coli cysteinyl-tRNA synthetase (cysS) was isolated by complementation of a strain deficient in cysteinyl-tRNA synthetase activity at high temperature (43 degrees C). Sequencing of a 2.1 kbp DNA fragment revealed an open reading frame of 1383 bp coding for a protein of 461 amino acid residues with a Mr of 52,280, a value in close agreement with that observed for the purified protein, which behaves as a monomer. The sequence of CysRS bears the canonical His-Ile- Gly -His (HIGH) and Lys-Met-Ser-Lys-Ser (KMSKS) motifs characteristic of the group of enzymes containing a Rossmann fold; furthermore, it shows striking homologies with MetRS (an homodimer of 677 residues) and to a lesser extent with Ile-, Leu-, and ValRS (monomers of 939, 860, and 951 residues respectively). With its monomeric state and smaller size, CysRS is probably more closely related to the primordial aminoacyl-tRNA synthetase from which all have diverged. Images PMID:2014166

  10. Momentum space orthogonal polynomial projection quantization

    NASA Astrophysics Data System (ADS)

    Handy, C. R.; Vrinceanu, D.; Marth, C. B.; Gupta, R.

    2016-04-01

    The orthogonal polynomial projection quantization (OPPQ) is an algebraic method for solving Schrödinger’s equation by representing the wave function as an expansion {{\\Psi }}(x)={\\displaystyle \\sum }n{{{Ω }}}n{P}n(x)R(x) in terms of polynomials {P}n(x) orthogonal with respect to a suitable reference function R(x), which decays asymptotically not faster than the bound state wave function. The expansion coefficients {{{Ω }}}n are obtained as linear combinations of power moments {μ }{{p}}=\\int {x}p{{\\Psi }}(x) {{d}}x. In turn, the {μ }{{p}}'s are generated by a linear recursion relation derived from Schrödinger’s equation from an initial set of low order moments. It can be readily argued that for square integrable wave functions representing physical states {{lim}}n\\to ∞ {{{Ω }}}n=0. Rapidly converging discrete energies are obtained by setting Ω coefficients to zero at arbitrarily high order. This paper introduces an extention of OPPQ in momentum space by using the representation {{Φ }}(k)={\\displaystyle \\sum }n{{{\\Xi }}}n{Q}n(k)T(k), where Q n (k) are polynomials orthogonal with respect to a suitable reference function T(k). The advantage of this new representation is that it can help solving problems for which there is no coordinate space moment equation. This is because the power moments in momentum space are the Taylor expansion coefficients, which are recursively calculated via Schrödinger’s equation. We show the convergence of this new method for the sextic anharmonic oscillator and an algebraic treatment of Gross-Pitaevskii nonlinear equation.

  11. Heisenberg algebra, umbral calculus and orthogonal polynomials

    SciTech Connect

    Dattoli, G.; Levi, D.; Winternitz, P.

    2008-05-15

    Umbral calculus can be viewed as an abstract theory of the Heisenberg commutation relation [P,M]=1. In ordinary quantum mechanics, P is the derivative and M the coordinate operator. Here, we shall realize P as a second order differential operator and M as a first order integral one. We show that this makes it possible to solve large classes of differential and integrodifferential equations and to introduce new classes of orthogonal polynomials, related to Laguerre polynomials. These polynomials are particularly well suited for describing the so-called flatenned beams in laser theory.

  12. Observations on the Proper Orthogonal Decomposition

    NASA Technical Reports Server (NTRS)

    Berkooz, Gal

    1992-01-01

    The Proper Orthogonal Decomposition (P.O.D.), also known as the Karhunen-Loeve expansion, is a procedure for decomposing a stochastic field in an L(2) optimal sense. It is used in diverse disciplines from image processing to turbulence. Recently the P.O.D. is receiving much attention as a tool for studying dynamics of systems in infinite dimensional space. This paper reviews the mathematical fundamentals of this theory. Also included are results on the span of the eigenfunction basis, a geometric corollary due to Chebyshev's inequality and a relation between the P.O.D. symmetry and ergodicity.

  13. Analysis of chromatograph systems using orthogonal collocation

    NASA Technical Reports Server (NTRS)

    Woodrow, P. T.

    1974-01-01

    Research is generating fundamental engineering design techniques and concepts for the chromatographic separator of a chemical analysis system for an unmanned, Martian roving vehicle. A chromatograph model is developed which incorporates previously neglected transport mechanisms. The numerical technique of orthogonal collocation is studied. To establish the utility of the method, three models of increasing complexity are considered, the latter two being limiting cases of the derived model: (1) a simple, diffusion-convection model; (2) a rate of adsorption limited, inter-intraparticle model; and (3) an inter-intraparticle model with negligible mass transfer resistance.

  14. 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.

  15. 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.

  16. Archaeal-type lysyl-tRNA synthetase in the Lyme disease spirochete Borrelia burgdorferi

    PubMed Central

    Ibba, Michael; Bono, James L.; Rosa, Patricia A.; Söll, Dieter

    1997-01-01

    Lysyl-tRNAs are essential for protein biosynthesis by ribosomal mRNA translation in all organisms. They are synthesized by lysyl-tRNA synthetases (EC 6.1.1.6), a group of enzymes composed of two unrelated families. In bacteria and eukarya, all known lysyl-tRNA synthetases are subclass IIc-type aminoacyl-tRNA synthetases, whereas some archaea have been shown to contain an unrelated class I-type lysyl-tRNA synthetase. Examination of the preliminary genomic sequence of the bacterial pathogen Borrelia burgdorferi, the causative agent of Lyme disease, indicated the presence of an open reading frame with over 55% similarity at the amino acid level to archaeal class I-type lysyl-tRNA synthetases. In contrast, no coding region with significant similarity to any class II-type lysyl-tRNA synthetase could be detected. Heterologous expression of this open reading frame in Escherichia coli led to the production of a protein with canonical lysyl-tRNA synthetase activity in vitro. Analysis of B. burgdorferi mRNA showed that the lysyl-tRNA synthetase-encoding gene is highly expressed, confirming that B. burgdorferi contains a functional class I-type lysyl-tRNA synthetase. The detection of an archaeal-type lysyl-tRNA synthetase in B. burgdorferi and other pathogenic spirochetes, but not to date elsewhere in bacteria or eukarya, indicates that the gene that encodes this enzyme has a common origin with its orthologue from the archaeal kingdom. This difference between the lysyl-tRNA synthetases of spirochetes and their hosts may be readily exploitable for the development of anti-spirochete therapeutics. PMID:9405621

  17. Geometric reconstruction of biological orthogonal plywoods.

    PubMed

    Aguilar Gutierrez, Oscar F; Rey, Alejandro D

    2016-01-28

    In this paper we focus on the structural determination of biological orthogonal plywoods, fiber-like composite analogues of liquid crystalline phases, where the fibrils of the building blocks show sharp 90° orientation jumps between fibers in adjacent domains. We present an original geometric and computational modelling that allows us to determine the fibrillary orientation in biological plywoods from periodic herringbone patterns commonly observed in cross-sections. Although herringbone patterns were long reported, the specific and quantitative relationships between herringbones and the orthogonal plywoods were absent or at best incomplete. Here we provide an efficient and new procedure to perform an inverse problem that connects two specific features of the herringbone patterns (aperture angle and wavelength) with the 3D morphology of the structure, whose accuracy and validity were ascertained through in silico simulations and also with real specimens ("Eremosphaera viridis"). This contribution extends significantly the better known characterization methods of 2D cross sections, such as the arced patterns observed in biological helicoidal plywoods, and with the present proposed methodology it adds another characterization tool for a variety of biological fibrous composites that form cornea-like tissues. PMID:26583687

  18. Response of transgenic poplar overexpressing cytosolic glutamine synthetase to phosphinothricin.

    PubMed

    Pascual, María Belén; Jing, Zhong Ping; Kirby, Edward G; Cánovas, Francisco M; Gallardo, Fernando

    2008-01-01

    Glutamine synthetase (GS) is the main enzyme involved in ammonia assimilation in plants and is the target of phosphinothricin (PPT), an herbicide commonly used for weed control in agriculture. As a result of the inhibition of GS, PPT also blocks photorespiration, resulting in the depletion of leaf amino acid pools leading to the plant death. Hybrid transgenic poplar (Populus tremula x P. alba INRA clone 7171-B4) overexpressing cytosolic GS is characterized by enhanced vegetative growth [Gallardo, F., Fu, J., Cantón, F.R., García-Gutiérrez, A., Cánovas, F.M., Kirby, E.G., 1999. Expression of a conifer glutamine synthetase gene in transgenic poplar. Planta 210, 19-26; Fu, J., Sampalo, R., Gallardo, F., Cánovas, F.M., Kirby, E.G., 2003. Assembly of a cytosolic pine glutamine synthetase holoenzyme in leaves of transgenic poplar leads to enhanced vegetative growth in young plants. Plant Cell Environ. 26, 411-418; Jing, Z.P., Gallardo, F., Pascual, M.B., Sampalo, R., Romero, J., Torres de Navarra, A., Cánovas, F.M., 2004. Improved growth in a field trial of transgenic hybrid poplar overexpressing glutamine synthetase. New Phytol. 164, 137-145], increased photosynthetic and photorespiratory capacities [El-Khatib, R.T., Hamerlynck, E.P., Gallardo, F., Kirby, E.G., 2004. Transgenic poplar characterized by ectopic expression of a pine cytosolic glutamine synthetase gene exhibits enhanced tolerance to water stress. Tree Physiol. 24, 729-736], enhanced tolerance to water stress (El-Khatib et al., 2004), and enhanced nitrogen use efficiency [Man, H.-M., Boriel, R., El-Khatib, R.T., Kirby, E.G., 2005. Characterization of transgenic poplar with ectopic expression of pine cytosolic glutamine synthetase under conditions of varying nitrogen availability. New Phytol. 167, 31-39]. In vitro plantlets of GS transgenic poplar exhibited enhanced resistance to PPT when compared with non-transgenic controls. After 30 days exposure to PPT at an equivalent dose of 275 g ha(-1), growth

  19. DOES IRON OR HEME CONTROL RAT HEPATIC DELTA-AMINOLEVULINIC ACID SYNTHETASE ACTIVITY

    EPA Science Inventory

    Disodium ethylenediamine tetraacetic acid and/or allylisopropylacetamide administration to rat pups did not evoke a premature induction of hepatic d-aminolevulinic acid synthetase. Administration of iron to adult rats did not alter d-aminolevulinic acid synthetase activity and ha...

  20. Bacteriophage T4 Virion Baseplate Thymidylate Synthetase and Dihydrofolate Reductase

    PubMed Central

    Kozloff, L. M.; Lute, M.; Crosby, L. K.

    1977-01-01

    Additional evidence is presented that both the phage T4D-induced thymidylate synthetase (gp td) and the T4D-induced dihydrofolate reductase (gp frd) are baseplate structural components. With regard to phage td it has been found that: (i) low levels of thymidylate synthetase activity were present in highly purified preparations of T4D ghost particles produced after infection with td+, whereas particles produced after infection with td− had no measurable enzymatic activity; (ii) a mutation of the T4D td gene from tdts to td+ simultaneously produced a heat-stable thymidylate synthetase enzyme and heat-stable phage particles (it should be noted that the phage baseplate structure determines heat lability); (iii) a recombinant of two T4D mutants constructed containing both tdts and frdts genes produced particles whose physical properties indicate that these two molecules physically interact in the baseplate. With regard to phage frd it has been found that two spontaneous revertants each of two different T4D frdts mutants to frd+ not only produced altered dihydrofolate reductases but also formed phage particles with heat sensitivities different from their parents. Properties of T4D particles produced after infection with parental T4D mutants presumed to have a deletion of the td gene and/or the frd gene indicate that these particles still retain some characteristics associated with the presence of both the td and the frd molecules. Furthermore, the particles produced by the deletion mutants have been found to be physically different from the parent particles. PMID:894793

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

    PubMed

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

    1992-10-01

    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

  2. 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

  3. How orthogonal are the OBO Foundry ontologies?

    PubMed Central

    2011-01-01

    Background Ontologies in biomedicine facilitate information integration, data exchange, search and query of biomedical data, and other critical knowledge-intensive tasks. The OBO Foundry is a collaborative effort to establish a set of principles for ontology development with the eventual goal of creating a set of interoperable reference ontologies in the domain of biomedicine. One of the key requirements to achieve this goal is to ensure that ontology developers reuse term definitions that others have already created rather than create their own definitions, thereby making the ontologies orthogonal. Methods We used a simple lexical algorithm to analyze the extent to which the set of OBO Foundry candidate ontologies identified from September 2009 to September 2010 conforms to this vision. Specifically, we analyzed (1) the level of explicit term reuse in this set of ontologies, (2) the level of overlap, where two ontologies define similar terms independently, and (3) how the levels of reuse and overlap changed during the course of this year. Results We found that 30% of the ontologies reuse terms from other Foundry candidates and 96% of the candidate ontologies contain terms that overlap with terms from the other ontologies. We found that while term reuse increased among the ontologies between September 2009 and September 2010, the level of overlap among the ontologies remained relatively constant. Additionally, we analyzed the six ontologies announced as OBO Foundry members on March 5, 2010, and identified that the level of overlap was extremely low, but, notably, so was the level of term reuse. Conclusions We have created a prototype web application that allows OBO Foundry ontology developers to see which classes from their ontologies overlap with classes from other ontologies in the OBO Foundry (http://obomap.bioontology.org). From our analysis, we conclude that while the OBO Foundry has made significant progress toward orthogonality during the period of this

  4. Inhibition of Pneumocystis carinii dihydropteroate synthetase by sulfa drugs.

    PubMed Central

    Merali, S; Zhang, Y; Sloan, D; Meshnick, S

    1990-01-01

    A new reversed-phase high-pressure liquid chromatography assay procedure for dihydropteroate synthetase (DHPS) that involves the elution of the enzyme incubation solution with a series of three solvents of decreasing polarity (ammonium phosphate buffer, 10% methanol, and 50% methanol) was designed. By this procedure DHPS was detected in Escherichia coli and Pneumocystis carinii with specific activities of 450 and 14 U/mg, respectively. A comparison of the effects of five sulfa drugs on P. carinii DHPS activity revealed that dapsone is the most potent of these drugs. PMID:2203302

  5. Beta-integrals and finite orthogonal systems of Wilson polynomials

    SciTech Connect

    Neretin, Yu A

    2002-08-31

    The integral is calculated and the system of orthogonal polynomials with weight equal to the corresponding integrand is constructed. This weight decreases polynomially, therefore only finitely many of its moments converge. As a result the system of orthogonal polynomials is finite. Systems of orthogonal polynomials related to {sub 5}H{sub 5}-Dougall's formula and the Askey integral is also constructed. All the three systems consist of Wilson polynomials outside the domain of positiveness of the usual weight.

  6. Orthogonal canonical forms for second-order systems

    NASA Technical Reports Server (NTRS)

    Williams, Trevor; Laub, Alan J.

    1992-01-01

    It is shown that a linear second-order system with arbitrary damping cannot be reduced to Hessenberg-triangular form by means of orthogonal transformations. However, it is also shown that such an orthogonal reduction is always possible for the modal damping commonly assumed for models of flexible structures. It is shown that modally damped models can be orthogonally reduced to a new triangular second-order Schur form.

  7. 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. PMID:26472323

  8. Multifunctional Surface Manipulation Using Orthogonal Click Chemistry.

    PubMed

    Brooks, Karson; Yatvin, Jeremy; McNitt, Christopher D; Reese, R Alexander; Jung, Calvin; Popik, Vladimir V; Locklin, Jason

    2016-07-01

    Polymer brushes are excellent substrates for the covalent immobilization of a wide variety of molecules due to their unique physicochemical properties and high functional group density. By using reactive microcapillary printing, poly(pentafluorophenyl acrylate) brushes with rapid kinetic rates toward aminolysis can be partially patterned with other click functionalities such as strained cyclooctyne derivatives and sulfonyl fluorides. This trireactive surface can then react locally and selectively in a one pot reaction via three orthogonal chemistries at room temperature: activated ester aminolysis, strain promoted azide-alkyne cycloaddition, and sulfur(VI) fluoride exchange, all of which are tolerant of ambient moisture and oxygen. Furthermore, we demonstrate that these reactions can also be used to create areas of morphologically distinct surface features on the nanoscale, by inducing buckling instabilities in the films and the grafting of nanoparticles. This approach is modular, and allows for the development of highly complex surface motifs patterned with different chemistry and morphology. PMID:27280689

  9. Confocal imaging with orthogonally polarized illumination beams

    NASA Astrophysics Data System (ADS)

    Kalita, Ranjan; Boruah, Bosanta R.

    2016-03-01

    In confocal microscopy the polarization of the illumination beam plays an important role in determining the orientation of the fluorescent molecules being illuminated. The efficiency of the excitation depends on the angle between the excitation electric field and the direction of the molecular dipole. In order to determine the orientation of the fluorescent molecules in the focal plane the molecules are to be excited using two mutually orthogonal electric fields. In this paper we show how a computer generated holography technique can be implemented using a ferroelectric liquid crystal spatial light modulator to conveniently obtain two images of the same target once with an X polarized illumination beam and another with a Y polarized illumination beam.

  10. 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.

  11. Description of sunspot cycles by orthogonal functions

    NASA Astrophysics Data System (ADS)

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

    1984-10-01

    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).

  12. 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).

  13. Orthogonal polynomials for refinable linear functionals

    NASA Astrophysics Data System (ADS)

    Laurie, Dirk; de Villiers, Johan

    2006-12-01

    A refinable linear functional is one that can be expressed as a convex combination and defined by a finite number of mask coefficients of certain stretched and shifted replicas of itself. The notion generalizes an integral weighted by a refinable function. The key to calculating a Gaussian quadrature formula for such a functional is to find the three-term recursion coefficients for the polynomials orthogonal with respect to that functional. We show how to obtain the recursion coefficients by using only the mask coefficients, and without the aid of modified moments. Our result implies the existence of the corresponding refinable functional whenever the mask coefficients are nonnegative, even when the same mask does not define a refinable function. The algorithm requires O(n^2) rational operations and, thus, can in principle deliver exact results. Numerical evidence suggests that it is also effective in floating-point arithmetic.

  14. 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. PMID:24921137

  15. 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.

  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. 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.

  18. 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

  19. LOCC indistinguishable orthogonal product quantum states.

    PubMed

    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

  20. 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. PMID:3121322

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

    PubMed Central

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

    1995-01-01

    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. PMID:7486915

  2. 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. PMID:18974016

  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. 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

  5. Holocarboxylase synthetase deficiency pre and post newborn screening

    PubMed Central

    Donti, Taraka R.; Blackburn, Patrick R.; Atwal, Paldeep S.

    2016-01-01

    Holocarboxylase synthetase deficiency is an autosomal recessive disorder of biotin metabolism resulting in multiple carboxylase deficiency. The typical presentation described in the medical literature is of neonatal onset within hours to weeks of birth with emesis, hypotonia, lethargy, seizures, metabolic ketolactic acidosis, hyperammonemia, developmental delay, skin rash and alopecia. The condition is screened for by newborn screening (NBS) tandem mass spectroscopy by elevated hydroxypentanoylcarnitine on dried blood spots. Urine organic acid profile may demonstrate elevated lactic, 3-OH isovaleric, 3-OH propionic, 3-MCC, methylcitric acids, and tiglylglycine consistent with loss of function of the above carboxylases. Here we describe a cohort of patients, 2 diagnosed pre-NBS and 3 post-NBS with broad differences in initial presentation and phenotype. In addition, prior to the advent of NBS, there are isolated reports of late-onset holocarboxylase synthetase deficiency in the medical literature, which describe patients diagnosed between 1 and 8 years of life, however to our knowledge there are no reports of late-onset HCLS being missed by NBS. Also we report two cases, each with novel pathogenic variants HCLS, diagnosed at age 3 years and 21 months respectively. The first patient had a normal newborn screen whilst the second had an abnormal newborn screen but was misdiagnosed as 3-methylcrotonylcarboxylase (3-MCC) deficiency and subsequently lost to follow-up until they presented again with severe metabolic acidosis. PMID:27114915

  6. Holocarboxylase synthetase deficiency pre and post newborn screening.

    PubMed

    Donti, Taraka R; Blackburn, Patrick R; Atwal, Paldeep S

    2016-06-01

    Holocarboxylase synthetase deficiency is an autosomal recessive disorder of biotin metabolism resulting in multiple carboxylase deficiency. The typical presentation described in the medical literature is of neonatal onset within hours to weeks of birth with emesis, hypotonia, lethargy, seizures, metabolic ketolactic acidosis, hyperammonemia, developmental delay, skin rash and alopecia. The condition is screened for by newborn screening (NBS) tandem mass spectroscopy by elevated hydroxypentanoylcarnitine on dried blood spots. Urine organic acid profile may demonstrate elevated lactic, 3-OH isovaleric, 3-OH propionic, 3-MCC, methylcitric acids, and tiglylglycine consistent with loss of function of the above carboxylases. Here we describe a cohort of patients, 2 diagnosed pre-NBS and 3 post-NBS with broad differences in initial presentation and phenotype. In addition, prior to the advent of NBS, there are isolated reports of late-onset holocarboxylase synthetase deficiency in the medical literature, which describe patients diagnosed between 1 and 8 years of life, however to our knowledge there are no reports of late-onset HCLS being missed by NBS. Also we report two cases, each with novel pathogenic variants HCLS, diagnosed at age 3 years and 21 months respectively. The first patient had a normal newborn screen whilst the second had an abnormal newborn screen but was misdiagnosed as 3-methylcrotonylcarboxylase (3-MCC) deficiency and subsequently lost to follow-up until they presented again with severe metabolic acidosis. PMID:27114915

  7. 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

  8. 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

  9. 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…

  10. A new method testing the orthogonality of different protecting groups.

    PubMed

    Ágoston, Károly; Ágoston, Ágnes; Dorgan, Colin R; Fügedi, Péter

    2015-12-11

    A new test was elaborated to identify a new set of orthogonal protecting groups. With the developed method eight different protecting groups were tested under various deprotection conditions and the complex reaction mixtures were analysed by HPLC. The developed method allows for quick identification of orthogonality using simple model structures. PMID:26580711

  11. 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.

  12. Direct evidence for an acyl phosphate intermediate in the folylpoly-. gamma. -glutamate synthetase and dihydrofolate synthetase-catalyzed reactions

    SciTech Connect

    Banerjee, R.

    1987-01-01

    The mechanism of the reactions catalyzed by two enzymes, namely dihydrofolate synthetase (DHFS) and folylpoly-..gamma..-glutamate synthetase (FPGS), has been investigated. The nature of the intermediate in each of the two reactions was monitored simultaneously in the multifunctional enzyme, FPGS/DHFS from E. coli. The latter was isolated from a transformant containing the cloned FPGS/DHFS gene. Incubation of (/sup 18/O)-H/sub 2/Pte and (/sup 17/O)-glutamate with ATP and the enzyme, resulted in the formation of (/sup 18/O)- and (/sup 17/O)-P/sub i/, thus providing strong evidence for the formation of an acyl phosphate species during catalysis of each reaction. The inorganic phosphate formed in the enzyme-catalyzed reaction was purified by chromatography on DEAE-cellulose, then converted to the trimethyl ester and analyzed by mass spectroscopy /sup 17/O NMR and /sup 31/P NMR. Stoichiometric formation of (/sup 17/O)- and (/sup 18/O)-Pi was observed. /sup 31/P NMR analysis showed the expected /sup 18/O-induced isotopic perturbations. The presence of (/sup 17/O)-trimethyl phosphate was revealed by /sup 17/O NMR. The mechanism of the FPGS-catalyzed reaction was also investigated with the antifolate (/sup 18/O)-methotrexate.

  13. Aminoacyl-tRNA synthetases as drug targets in eukaryotic parasites☆

    PubMed Central

    Pham, James S.; Dawson, Karen L.; Jackson, Katherine E.; Lim, Erin E.; Pasaje, Charisse Flerida A.; Turner, Kelsey E.C.; Ralph, Stuart A.

    2013-01-01

    Aminoacyl-tRNA synthetases are central enzymes in protein translation, providing the charged tRNAs needed for appropriate construction of peptide chains. These enzymes have long been pursued as drug targets in bacteria and fungi, but the past decade has seen considerable research on aminoacyl-tRNA synthetases in eukaryotic parasites. Existing inhibitors of bacterial tRNA synthetases have been adapted for parasite use, novel inhibitors have been developed against parasite enzymes, and tRNA synthetases have been identified as the targets for compounds in use or development as antiparasitic drugs. Crystal structures have now been solved for many parasite tRNA synthetases, and opportunities for selective inhibition are becoming apparent. For different biological reasons, tRNA synthetases appear to be promising drug targets against parasites as diverse as Plasmodium (causative agent of malaria), Brugia (causative agent of lymphatic filariasis), and Trypanosoma (causative agents of Chagas disease and human African trypanosomiasis). Here we review recent developments in drug discovery and target characterisation for parasite aminoacyl-tRNA synthetases. PMID:24596663

  14. 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. PMID:19088795

  15. 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.

  16. Sequence, structural and evolutionary relationships between class 2 aminoacyl-tRNA synthetases.

    PubMed Central

    Cusack, S; Härtlein, M; Leberman, R

    1991-01-01

    Class 2 aminoacyl-tRNA synthetases, which include the enzymes for alanine, aspartic acid, asparagine, glycine, histidine, lysine, phenylalanine, proline, serine and threonine, are characterised by three distinct sequence motifs 1,2 and 3 (reference 1). The structural and evolutionary relatedness of these ten enzymes are examined using alignments of primary sequences from prokaryotic and eukaryotic sources and the known three dimensional structure of seryl-tRNA synthetase from E. coli. It is shown that motif 1 forms part of the dimer interface of seryl-tRNA synthetase and motifs 2 and 3 part of the putative active site. It is further shown that the seven alpha 2 dimeric synthetases can be subdivided into class 2a (proline, threonine, histidine and serine) and class 2b (aspartic acid, asparagine and lysine), each subclass sharing several important characteristic sequence motifs in addition to those characteristic of class 2 enzymes in general. The alpha 2 beta 2 tetrameric enzymes (for glycine and phenylalanine) show certain special features in common as well as some of the class 2b motifs. In the alanyl-tRNA synthetase only motif 3 and possibly motif 2 can be identified. The sequence alignments suggest that the catalytic domain of other class 2 synthetases should resemble the antiparallel domain found in seryl-tRNA synthetase. Predictions are made about the sequence location of certain important helices and beta-strands in this domain as well as suggestions concerning which residues are important in ATP and amino acid binding. Strong homologies are found in the N-terminal extensions of class 2b synthetases and in the C-terminal extensions of class 2a synthetases suggesting that these putative tRNA binding domains have been added at a later stage in evolution to the catalytic domain. Images PMID:1852601

  17. Regulation of β-Glucan Synthetase Activity by Auxin in Pea Stem Tissue

    PubMed Central

    Ray, Peter M.

    1973-01-01

    Treatment of pea stem segments with indoleacetic acid (IAA) causes within 1 hour a 2- to 4-fold increase in activity of particulate uridine diphosphoglucose-dependent β-glucan synthetase obtainable from the tissue. The IAA effect is observable in tissue from all parts of the elongation zone of the pea stem, and also in older tissue that is not capable of a cell enlargement response to IAA. A large increase in activity is caused by IAA only if synthetase activity in the isolated tissue has first been allowed to fall substantially below the intact plant level, and only if sucrose is supplied along with IAA. Treatment of tissue with sucrose alone after a period of sugar starvation causes a transient rise of synthetase activity. The decline in synthetase activity in absence of IAA, the rise caused by IAA, and the transient rise caused by sucrose are all strongly temperature-dependent. IAA and sucrose do not affect the activity of isolated synthetase particles. Synthetase activity in vivo is sensitive to as low as 0.1 μm IAA and is increased by IAA analogues that are active as auxins on elongation but not by nonauxin analogues. Activity begins to rise 10 to 15 minutes after exposure to IAA, which places this among the most rapid enzyme effects of a plant growth regulator heretofore demonstrated, and among the most rapid known metabolic effects of auxins. The effect is seen also with polysaccharide synthetase activity using uridine diphosphate-galactose or uridine diphosphate-xylose as substrates, and to a lesser extent with guanosine diphosphoglucose-dependent glucan synthetase activity. Glucan synthetase from IAA-treated tissue appears to have a higher affinity for uridine diphosphate-glucose than the control. PMID:16658379

  18. 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. PMID:25955995

  19. Directional lapped orthogonal transform: theory and design.

    PubMed

    Muramatsu, Shogo; Han, Dandan; Kobayashi, Tomoya; Kikuchi, Hisakazu

    2012-05-01

    This paper proposes a directional design method of 2-D nonseparable linear-phase paraunitary filter banks. The proposed method is based on a lattice structure consisting of the 2-D separable DCT block and nonseparable support extension processes. Because of the nonseparability, the bases are allowed to be directional with the critically fixed subsampling, overlapping, orthogonal, symmetric, real-valued, and compact support properties. First, a novel vanishing moment (VM) condition is introduced as a suitable directional constraint, where the moment is referred to as the trend VM. The condition forces wavelet filters, i.e., high-pass and bandpass filters, to annihilate trend-surface components. Second, some theoretical properties of TVMs are discussed for general 2-D paraunitary systems, and then, the properties are applied to the lattice parameters. In order to verify the significance, several design examples are shown, the trend-surface annihilation properties are numerically confirmed, and the denoising capability is evaluated for images through shrinkage. It is shown that our proposed transforms yield perceptually preferable results. PMID:22231175

  20. 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.

  1. The Representation and Parametrization of Orthogonal Matrices.

    PubMed

    Shepard, Ron; Brozell, Scott R; Gidofalvi, Gergely

    2015-07-16

    Four representations and parametrizations of orthogonal matrices Q ∈ R(m×n) in terms of the minimal number of essential parameters {φ} are discussed: the exponential representation, the Householder reflector representation, the Givens rotation representation, and the rational Cayley transform representation. Both square n = m and rectangular n < m situations are considered. Two separate kinds of parametrizations are considered: one in which the individual columns of Q are distinct, the Stiefel manifold, and the other in which only span(Q) is significant, the Grassmann manifold. The practical issues of numerical stability, continuity, and uniqueness are discussed. The computation of Q in terms of the essential parameters {φ}, and also the extraction of {φ} for a given Q are considered for all of the parametrizations. The transformation of gradient arrays between the Q and {φ} variables is discussed for all representations. It is our hope that developers of new methods will benefit from this comparative presentation of an important but rarely analyzed subject. PMID:25946418

  2. Genuinely multipartite entangled states and orthogonal arrays

    NASA Astrophysics Data System (ADS)

    Goyeneche, Dardo; Życzkowski, Karol

    2014-08-01

    A pure quantum state of N subsystems with d levels each is called k-multipartite maximally entangled state, which we call a k-uniform state, if all its reductions to k qudits are maximally mixed. These states form a natural generalization of N-qudit Greenberger-Horne-Zeilinger states which belong to the class 1-uniform states. We establish a link between the combinatorial notion of orthogonal arrays and k-uniform states and prove the existence of several classes of such states for N-qudit systems. In particular, known Hadamard matrices allow us to explicitly construct 2-uniform states for an arbitrary number of N >5 qubits. We show that finding a different class of 2-uniform states would imply the Hadamard conjecture, so the full classification of 2-uniform states seems to be currently out of reach. Furthermore, we establish links between the existence of k-uniform states and classical and quantum error correction codes and provide a graph representation for such states.

  3. 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.

  4. 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.

  5. Quantum nonlocality of multipartite orthogonal product states

    NASA Astrophysics Data System (ADS)

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

    2016-03-01

    Local distinguishability of orthogonal quantum states is an area of active research in quantum information theory. However, most of the relevant results are about local distinguishability in bipartite Hilbert space and very little is known about the multipartite case. In this paper we present a generic method to construct a completable n -partite (n ≥3 ) product basis with only 2 n members, which exhibits nonlocality without entanglement with n parties, each holding a system of any finite dimension. We give an effective proof of the nonlocality of the completable multipartite product basis. In addition, we construct another incomplete multipartite product basis with a smaller number of members that cannot be distinguished by local operations and classical communication in a d1⊗d2⊗⋯⊗dn quantum system, where n ≥3 and di≥2 for i =1 ,2 ,...,n . The results can lead to a better understanding of the phenomenon of nonlocality without entanglement in any multipartite quantum system.

  6. Properties of Kaurene Synthetase from Marah macrocarpus1

    PubMed Central

    Frost, Russell G.; West, Charles A.

    1977-01-01

    The kaurene synthetase from immature seeds of Marah macrocarpus (Greene) Greene was partially purified from cell-free homogenates of endosperm by a combination of QAE-Sephadex A-25 chromatography and hydroxyapatite chromatography and freed of contaminating phosphatase activity. The two catalytic activities associated with kaurene synthetase, the cyclization of geranylgeranyl-pyrophosphate to copalyl-pyrophosphate (activity A) and the cyclization of copalyl-pyrophosphate to ent-kaurene (activity B), were not even partially resolved from one another during these procedures. Both activities had identical elution profiles from a calibrated Sepharose 4B column corresponding to a molecular weight less than that of ovalbumin (45,000). The A and B activities had pH optima of 7.3 and 6.9, respectively. Both activities required millimolar concentrations of the following divalent cations in the order: Mg2+ > Mn2+ > Co2+. Activities A and B were both sensitive to inhibition by Hg2+, Cu2+, p-hydroxymercuribenzoate, and N-ethylmaleimide, but activity B was much more sensitive than activity A. The average value of Km′ (apparent Km in the absence of substrate inhibition) for geranylgeranyl-pyrophosphate was 1.6 μm. Values of 0.5 and 0.6 μm were obtained for Km′ and Km, respectively, for copalyl-pyrophosphate. The Vm′ values for the two activities were similar: 12 and 9 pmol/minute·μg protein for activities A and B, respectively. N,N-Dimethylaminoethyl-2,2-diphenylpentanoate (SKF-525A) and N,N-dimethylaminoethyl-2,2-diphenylphentyl ether (SKF-3301A), tributyl-2,4-dichlorobenzylphosphonium chloride (Phosfon D), tributyl-2,4-dichlorobenzylammonium chloride (Phosfon S), 2′-isopropyl-4′-(trimethylammonium chloride)-5′-methylphenyl piperidine-1-carboxylate (Amo-1618), 2-(N,N-dimethyl-N-heptylammonium bromide)-p-methan-1-ol (Q-58), and 2-(N,N-dimethyl-N-octylammonium bromide)-p-methan-1-ol (Q-64), at concentrations from 1 to 5 μm, were effective inhibitors of kaurene

  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. Altering the Enantioselectivity of Tyrosyl-tRNA Synthetase by Insertion of a Stereospecific Editing Domain.

    PubMed

    Richardson, Charles J; First, Eric A

    2016-03-15

    Translation of mRNAs by the ribosome is stereospecific, with only l-amino acids being incorporated into the nascent polypeptide chain. This stereospecificity results from the exclusion of d-amino acids at three steps during protein synthesis: (1) the aminoacylation of tRNA by aminoacyl-tRNA synthetases, (2) binding of aminoacyl-tRNAs to EF-Tu, and (3) recognition of aminoacyl-tRNAs by the ribosome. As a first step toward incorporating d-amino acids during protein synthesis, we have altered the enantioselectivity of tyrosyl-tRNA synthetase. This enzyme is unusual among aminoacyl-tRNA synthetases, as it can aminoacylate tRNA with d-tyrosine (albeit at a reduced rate compared to l-tyrosine). To change the enantioselectivity of tyrosyl-tRNA synthetase, we introduced the post-transfer editing domain from Pyrococcus horikoshii phenylalanyl-tRNA synthetase into the connective polypeptide 1 (CP1) domain of Geobacillus stearothermophilus tyrosyl-tRNA synthetase (henceforth designated TyrRS-FRSed). We show that the phenylalanyl-tRNA synthetase editing domain is stereospecific, hydrolyzing l-Tyr-tRNA(Tyr), but not d-Tyr-tRNA(Tyr). We further show that inserting the phenylalanyl-tRNA synthetase editing domain into the CP1 domain of tyrosyl-tRNA synthetase decreases the activity of the synthetic site in tyrosyl-tRNA synthetase. This decrease in activity is critical, as it prevents the rate of synthesis from overwhelming the ability of the editing domain to hydrolyze the l-Tyr-tRNA(Tyr) product. Overall, inserting the phenylalanyl-tRNA synthetase editing domain results in a 2-fold shift in the enantioselectivity of tyrosyl-tRNA synthetase toward the d-Tyr-tRNA(Tyr) product. When a 4-fold excess of d-tyrosine is used, approximately 40% of the tRNA(Tyr) is aminoacylated with d-tyrosine. PMID:26890980

  9. Treatment of renal colic by prostaglandin synthetase inhibitors and avafortan (analgesic antispasmodic).

    PubMed

    el-Sherif, A E; Foda, R; Norlen, L J; Yahia, H

    1990-12-01

    In a study of the pain-relieving effect of 3 drugs commonly used to treat acute renal colic in this hospital, intravenous indomethacin and intramuscular diclofenac (prostaglandin synthetase inhibitors) were compared with intravenous Avafortan (analgesic antispasmodic). As first-line analgesics, prostaglandin synthetase inhibitors, if given intravenously, offer an effective alternative to Avafortan. Of 145 patients studied, 32 required a second injection for complete relief of pain. Administering a second dose of prostaglandin synthetase inhibitors resulted in equally significant pain relief rate even though the route was intramuscular. PMID:2265331

  10. Multiple molecular forms of glutamine synthetase in pea seeds.

    PubMed

    Antonyuk, L P; Pushkin, A V; Vorobyeva, L M; Solovjeva, N A; Evstigneeva, Z G; Kretovich, W L

    1982-08-20

    Multiple molecular forms of glutamine synthetase (GS, EC 6.3.1.2) have been studied in pea seeds of different varieties. The number of GS molecular forms in the seeds proved to be related to their colour. Two GS forms in the green seeds have been found and only one of them in the yellow seeds. Green seeds had chlorophyll content amounted to 0.4% of the total pigment content in the leaves. Chloroplasts, somewhat smaller than those in pea leaves of the same variety, have been isolated from green seeds. The presence of the second GS form in the pea green seeds we relate to the chloroplasts. By electrophoretic mobility both forms of GS from the green seeds are not identical to the chloroplast GS and the cytosol GS of leaves. Thus, we believe pea plant to contain, at least, four GS forms. PMID:6127624

  11. Aminoacyl-tRNA synthetase complexes: molecular multitasking revealed

    PubMed Central

    Hausmann, Corinne D.; Ibba, Michael

    2008-01-01

    The accurate synthesis of proteins, dictated by the corresponding nucleotide sequence encoded in mRNA, is essential for cell growth and survival. Central to this process are the aminoacyl-tRNA synthetases (aaRSs), which provide amino acid substrates for the growing polypeptide chain in the form of aminoacyl-tRNAs. The aaRSs are essential for coupling the correct amino acid and tRNA molecules, but are also known to associate in higher order complexes with proteins involved in processes beyond translation. Multiprotein complexes containing aaRSs are found in all three domains of life playing roles in splicing, apoptosis, viral assembly, and regulation of transcription and translation. An overview of the complexes aaRSs form in all domains of life is presented, demonstrating the extensive network of connections between the translational machinery and cellular components involved in a myriad of essential processes beyond protein synthesis. PMID:18522650

  12. S-adenosylmethionine synthetase in bloodstream Trypanosoma brucei.

    PubMed

    Yarlett, N; Garofalo, J; Goldberg, B; Ciminelli, M A; Ruggiero, V; Sufrin, J R; Bacchi, C J

    1993-03-24

    S-adenosylmethionine synthetase was studied from bloodstream forms of Trypanosoma brucei brucei, the agent of African sleeping sickness. Two isoforms of the enzyme were evident from Eadie Hofstee and Hanes-Woolf plots of varying ATP or methionine concentrations. In the range 10-250 microM the Km for methionine was 20 microM, and this changed to 200 microM for the range 0.5-5.0 mM. In the range 10-250 microM the Km for ATP was 53 microM, and this changed to 1.75 mM for the range 0.5-5.0 mM. The trypanosome enzyme had a molecular weight of 145 kDa determined by agarose gel filtration. Methionine analogs including selenomethionine, L-2-amino-4-methoxy-cis but-3-enoic acid and ethionine acted as competitive inhibitors of methionine and as weak substrates when tested in the absence of methionine with [14C]ATP. The enzyme was not inducible in procyclic trypomastigotes in vitro, and the enzyme half-life was > 6 h. T. b. brucei AdoMet synthetase was inhibited by AdoMet (Ki 240 microM). The relative insensitivity of the trypanosome enzyme to control by product inhibition indicates it is markedly different from mammalian isoforms of the enzyme which are highly sensitive to AdoMet. Since trypanosomes treated with the ornithine decarboxylase antagonist DL-alpha-difluoromethylornithine accumulate AdoMet and dcAdoMet (final concentration approximately 5 mM), this enzyme may be the critical drug target linking inhibition of polyamine synthesis to disruption of AdoMet metabolism. PMID:8457607

  13. 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)

  14. Cavitation as a mechanism of substrate discrimination by adenylosuccinate synthetases.

    PubMed

    Iancu, Cristina V; Zhou, Yang; Borza, Tudor; Fromm, Herbert J; Honzatko, Richard B

    2006-09-26

    Adenylosuccinate synthetase catalyzes the first committed step in the de novo biosynthesis of AMP, coupling L-aspartate and IMP to form adenylosuccinate. Km values of IMP and 2'-deoxy-IMP are nearly identical with each substrate supporting comparable maximal velocities. Nonetheless, the Km value for L-aspartate and the Ki value for hadacidin (a competitive inhibitor with respect to L-aspartate) are 29-57-fold lower in the presence of IMP than in the presence of 2'-deoxy-IMP. Crystal structures of the synthetase ligated with hadacidin, GDP, and either 6-phosphoryl-IMP or 2'-deoxy-6-phosphoryl-IMP are identical except for the presence of a cavity normally occupied by the 2'-hydroxyl group of IMP. In the presence of 6-phosphoryl-IMP and GDP (hadacidin absent), the L-aspartate pocket can retain its fully ligated conformation, forming hydrogen bonds between the 2'-hydroxyl group of IMP and sequence-invariant residues. In the presence of 2'-deoxy-6-phosphoryl-IMP and GDP, however, the L-aspartate pocket is poorly ordered. The absence of the 2'-hydroxyl group of the deoxyribonucleotide may destabilize binding of the ligand to the L-aspartate pocket by disrupting hydrogen bonds that maintain a favorable protein conformation and by the introduction of a cavity into the fully ligated active site. At an approximate energy cost of 2.2 kcal/mol, the unfavorable thermodynamics of cavity formation may be the major factor in destabilizing ligands at the L-aspartate pocket. PMID:16981730

  15. 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

  16. Ocean Models and Proper Orthogonal Decomposition

    NASA Astrophysics Data System (ADS)

    Salas-de-Leon, D. A.

    2007-05-01

    The increasing computational developments and the better understanding of mathematical and physical systems resulted in an increasing number of ocean models. Long time ago, modelers were like a secret organization and recognize each other by using secret codes and languages that only a select group of people was able to recognize and understand. The access to computational systems was reduced, on one hand equipment and the using time of computers were expensive and restricted, and on the other hand, they required an advance computational languages that not everybody wanted to learn. Now a days most college freshman own a personal computer (PC or laptop), and/or have access to more sophisticated computational systems than those available for research in the early 80's. The resource availability resulted in a mayor access to all kind models. Today computer speed and time and the algorithms does not seem to be a problem, even though some models take days to run in small computational systems. Almost every oceanographic institution has their own model, what is more, in the same institution from one office to the next there are different models for the same phenomena, developed by different research member, the results does not differ substantially since the equations are the same, and the solving algorithms are similar. The algorithms and the grids, constructed with algorithms, can be found in text books and/or over the internet. Every year more sophisticated models are constructed. The Proper Orthogonal Decomposition is a technique that allows the reduction of the number of variables to solve keeping the model properties, for which it can be a very useful tool in diminishing the processes that have to be solved using "small" computational systems, making sophisticated models available for a greater community.

  17. Perturbations around the zeros of classical orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Sasaki, Ryu

    2015-04-01

    Starting from degree N solutions of a time dependent Schrödinger-like equation for classical orthogonal polynomials, a linear matrix equation describing perturbations around the N zeros of the polynomial is derived. The matrix has remarkable Diophantine properties. Its eigenvalues are independent of the zeros. The corresponding eigenvectors provide the representations of the lower degree ( 0 , 1 , … , N - 1 ) polynomials in terms of the zeros of the degree N polynomial. The results are valid universally for all the classical orthogonal polynomials, including the Askey scheme of hypergeometric orthogonal polynomials and its q-analogues.

  18. 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. PMID:25697992

  19. 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.

  20. 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.

  1. Cloning and characterization of the gene for Escherichia coli seryl-tRNA synthetase.

    PubMed Central

    Härtlein, M; Madern, D; Leberman, R

    1987-01-01

    Seryl-tRNA synthetase is the gene product of the serS locus in Escherichia coli. Its gene has been cloned by complementation of a serS temperature sensitive mutant K28 with an E. coli gene bank DNA. The resulting clones overexpress seryl-tRNA synthetase by a factor greater than 50 and more than 6% of the total cellular protein corresponds to the enzyme. The DNA sequence of the complete coding region and the 5'- and 3' untranslated regions was determined. Protein sequence comparison of SerRS with all available aminoacyl-tRNA synthetase sequences revealed some regions of significant homology particularly with the isoleucyl- and phenylalanyl-tRNA synthetases from E. coli. Images PMID:3029694

  2. Mutational Separation of Aminoacylation and Cytokine Activities of Human Tyrosyl-tRNA Synthetase

    PubMed Central

    Kapoor, Mili; Otero, Francella J.; Slike, Bonnie M.; Ewalt, Karla L.; Yang, Xiang-Lei

    2009-01-01

    SUMMARY Aminoacyl-tRNA synthetases are known for catalysis of aminoacylation. Significantly, some mammalian synthetases developed cytokine functions possibly linked to disease-causing mutations in tRNA synthetases. Not understood is how epitopes for cytokine signaling were introduced into catalytic scaffolds without disturbing aminoacylation. Here we investigate human tyrosyl-tRNA synthetase, where a catalytic-domain surface helix—next to the active site—was recruited for IL-8-like cytokine signaling. Taking advantage of our high-resolution structure, the reciprocal impact of rational mutations designed to disrupt aminoacylation or cytokine signaling was investigated with multiple assays. The collective analysis demonstrated a protective fine–structure separation of aminoacylation from cytokine activities within the conserved catalytic domain. As a consequence, disease-causing mutations affecting cell signaling can arise without disturbing aminoacylation. These results with TyrRS also predict the previously unknown binding conformation of IL-8-like CXC cytokines. PMID:19477417

  3. Induction of angiogenesis by a fragment of human tyrosyl-tRNA synthetase.

    PubMed

    Wakasugi, Keisuke; Slike, Bonnie M; Hood, John; Ewalt, Karla L; Cheresh, David A; Schimmel, Paul

    2002-06-01

    The first step of protein synthesis is catalyzed by aminoacyl-tRNA synthetases. In addition, certain mammalian tRNA synthetases link protein synthesis to cytokine signaling pathways. In particular, human tyrosyl-tRNA synthetase (TyrRS) can be split by proteolysis into two fragments having distinct cytokine activities. One of the TyrRS fragments (mini TyrRS) contains features identical to those in CXC chemokines (like interleukin-8) that also act as angiogenic factors. Here mini TyrRS (but not full-length TyrRS) is shown to stimulate chemotaxis of endothelial cells in vitro and stimulate angiogenesis in each of two in vivo animal models. The angiogenic activity of mini TyrRS can be opposed by anti-angiogenic chemokines like IP-10. Thus, a biological fragment of human tyrosyl-tRNA synthetase links protein synthesis to regulation of angiogenesis. PMID:11956181

  4. Mutational separation of aminoacylation and cytokine activities of human tyrosyl-tRNA synthetase.

    PubMed

    Kapoor, Mili; Otero, Francella J; Slike, Bonnie M; Ewalt, Karla L; Yang, Xiang-Lei

    2009-05-29

    Aminoacyl tRNA synthetases are known for catalysis of aminoacylation. Significantly, some mammalian synthetases developed cytokine functions possibly linked to disease-causing mutations in tRNA synthetases. Not understood is how epitopes for cytokine signaling were introduced into catalytic scaffolds without disturbing aminoacylation. Here we investigate human tyrosyl-tRNA synthetase, where a catalytic-domain surface helix, next to the active site, was recruited for interleukin-8-like cytokine signaling. Taking advantage of our high resolution structure, the reciprocal impact of rational mutations designed to disrupt aminoacylation or cytokine signaling was investigated with multiple assays. The collective analysis demonstrated a protective fine-structure separation of aminoacylation from cytokine activities within the conserved catalytic domain. As a consequence, disease-causing mutations affecting cell signaling can arise without disturbing aminoacylation. These results with TyrRS also predict the previously unknown binding conformation of interleukin-8-like CXC cytokines. PMID:19477417

  5. Nodule-specific modulation of glutamine synthetase in transgenic Medicago truncatula leads to inverse alterations in asparagine synthetase expression.

    PubMed

    Carvalho, Helena G; Lopes-Cardoso, Inês A; Lima, Ligia M; Melo, Paula M; Cullimore, Julie V

    2003-09-01

    Transgenic Medicago truncatula plants were produced harboring chimeric gene constructs of the glutamine synthetase (GS) cDNA clones (MtGS1a or MtGS1b) fused in sense or antisense orientation to the nodule-specific leghemoglobin promoter Mtlb1. A series of transgenic plants were obtained showing a 2- to 4-fold alteration in nodule GS activity when compared with control plants. Western and northern analyses revealed that the increased or decreased levels of GS activity correlate with the amount of cytosolic GS polypeptides and transcripts present in the nodule extracts. An analysis of the isoenzyme composition showed that the increased or decreased levels of GS activity were attributable to major changes in the homo-octameric isoenzyme GS1a. Nodules of plants transformed with antisense GS constructs showed an increase in the levels of both asparagine synthetase (AS) polypeptides and transcripts when compared with untransformed control plants, whereas the sense GS transformants showed decreased AS transcript levels but polypeptide levels similar to control plants. The polypeptide abundance of other nitrogen metabolic enzymes NADH-glutamic acid synthase and aspartic acid amino-transferase as well as those of major carbon metabolic enzymes phosphoenolpyruvate carboxylase, carbonic anhydrase, and sucrose synthase were not affected by the GS-gene manipulations. Increased levels of AS polypeptides and transcripts were also transiently observed in nodules by inhibiting GS activity with phosphinothricin. Taken together, the results presented here suggest that GS activity negatively regulates the level of AS in root nodules of M. truncatula. The potential role of AS in assimilating ammonium when GS becomes limiting is discussed. PMID:12970490

  6. Nodule-Specific Modulation of Glutamine Synthetase in Transgenic Medicago truncatula Leads to Inverse Alterations in Asparagine Synthetase Expression1

    PubMed Central

    Carvalho, Helena G.; Lopes-Cardoso, Inês A.; Lima, Ligia M.; Melo, Paula M.; Cullimore, Julie V.

    2003-01-01

    Transgenic Medicago truncatula plants were produced harboring chimeric gene constructs of the glutamine synthetase (GS) cDNA clones (MtGS1a or MtGS1b) fused in sense or antisense orientation to the nodule-specific leghemoglobin promoter Mtlb1. A series of transgenic plants were obtained showing a 2- to 4-fold alteration in nodule GS activity when compared with control plants. Western and northern analyses revealed that the increased or decreased levels of GS activity correlate with the amount of cytosolic GS polypeptides and transcripts present in the nodule extracts. An analysis of the isoenzyme composition showed that the increased or decreased levels of GS activity were attributable to major changes in the homo-octameric isoenzyme GS1a. Nodules of plants transformed with antisense GS constructs showed an increase in the levels of both asparagine synthetase (AS) polypeptides and transcripts when compared with untransformed control plants, whereas the sense GS transformants showed decreased AS transcript levels but polypeptide levels similar to control plants. The polypeptide abundance of other nitrogen metabolic enzymes NADH-glutamic acid synthase and aspartic acid amino-transferase as well as those of major carbon metabolic enzymes phosphoenolpyruvate carboxylase, carbonic anhydrase, and sucrose synthase were not affected by the GS-gene manipulations. Increased levels of AS polypeptides and transcripts were also transiently observed in nodules by inhibiting GS activity with phosphinothricin. Taken together, the results presented here suggest that GS activity negatively regulates the level of AS in root nodules of M. truncatula. The potential role of AS in assimilating ammonium when GS becomes limiting is discussed. PMID:12970490

  7. 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.

  8. Genetic validation of aminoacyl-tRNA synthetases as drug targets in Trypanosoma brucei.

    PubMed

    Kalidas, Savitha; Cestari, Igor; Monnerat, Severine; Li, Qiong; Regmi, Sandesh; Hasle, Nicholas; Labaied, Mehdi; Parsons, Marilyn; Stuart, Kenneth; Phillips, Margaret A

    2014-04-01

    Human African trypanosomiasis (HAT) is an important public health threat in sub-Saharan Africa. Current drugs are unsatisfactory, and new drugs are being sought. Few validated enzyme targets are available to support drug discovery efforts, so our goal was to obtain essentiality data on genes with proven utility as drug targets. Aminoacyl-tRNA synthetases (aaRSs) are known drug targets for bacterial and fungal pathogens and are required for protein synthesis. Here we survey the essentiality of eight Trypanosoma brucei aaRSs by RNA interference (RNAi) gene expression knockdown, covering an enzyme from each major aaRS class: valyl-tRNA synthetase (ValRS) (class Ia), tryptophanyl-tRNA synthetase (TrpRS-1) (class Ib), arginyl-tRNA synthetase (ArgRS) (class Ic), glutamyl-tRNA synthetase (GluRS) (class 1c), threonyl-tRNA synthetase (ThrRS) (class IIa), asparaginyl-tRNA synthetase (AsnRS) (class IIb), and phenylalanyl-tRNA synthetase (α and β) (PheRS) (class IIc). Knockdown of mRNA encoding these enzymes in T. brucei mammalian stage parasites showed that all were essential for parasite growth and survival in vitro. The reduced expression resulted in growth, morphological, cell cycle, and DNA content abnormalities. ThrRS was characterized in greater detail, showing that the purified recombinant enzyme displayed ThrRS activity and that the protein localized to both the cytosol and mitochondrion. Borrelidin, a known inhibitor of ThrRS, was an inhibitor of T. brucei ThrRS and showed antitrypanosomal activity. The data show that aaRSs are essential for T. brucei survival and are likely to be excellent targets for drug discovery efforts. PMID:24562907

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

    PubMed Central

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

    2015-01-01

    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. PMID:25697791

  10. 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.

  11. Effect of heat shock on poly(ADP-ribose) synthetase and DNA repair in Drosophila cells

    SciTech Connect

    Nolan, N.L.; Kidwell, W.R.

    1982-04-01

    Poly(ADP-ribose) synthetase, a chromatin-bound enzyme which attaches polyanionic chains of ADP-ribose to nuclear proteins, was found to be temperature sensitive in intact Drosophila melanogaster cells. The synthetase was completely inactivated by heat-shocking the cells at 37/sup 0/C for 5 min, a condition which had no appreciable effect on the subsequent growth of Drosophila cells at their physiological temperature. The heat-shock effect on synthetase was reversible; enzyme activity began to reappear about 2 hr post heat shock. During the 2-hr interval when poly(ADP-ribose) synthetase was absent, the cells were competent in repair of ..gamma..-ray-induced DNA strand breaks as shown by DNA sedimentation studies on alkaline sucrose gradients. It is thus concluded that poly(ADP-ribose) synthesis is unnecessary for repair of DNA strand breaks introduced by irradiation. The same conclusion was reached from the fact that two inhibitors of poly(ADP-ribose) synthetase 3-aminobenzamide and 5-methylnicotinamide, failed to block repair of ..gamma..-ray-induced DNA chain breaks even though both inhibitors reduced the amount of poly(ADP-ribose) synthesized in cells by 50-75%. Although it was found that the repair of DNA strand breaks is independent of poly(ADP-ribose) synthesis, irradiation does activate the synthetase in control cells, as shown by radioimmunoassay of poly(ADP-ribose) levels.

  12. Gain-of-function mutational activation of human tRNA synthetase procytokine.

    PubMed

    Yang, Xiang-Lei; Kapoor, Mili; Otero, Francella J; Slike, Bonnie M; Tsuruta, Hiro; Frausto, Ricardo; Bates, Alison; Ewalt, Karla L; Cheresh, David A; Schimmel, Paul

    2007-12-01

    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. PMID:18096501

  13. Analysis of orthogonal waveform for spaceborne MIMO-GMTI radar

    NASA Astrophysics Data System (ADS)

    Zou, Bo; Dong, Zhen; Du, Xiang-yu

    2011-10-01

    The application of MIMO (Multiple input multiple output) techniques to spaceborne multichannel radar offers a number of advantages, including target detection, parameter estimation, and so on. Based on two kinds of waveforms presented in MIMO radar, a concise definition of synthetical ISLR is proposed. Through analysis of synthetical ISLR for two kinds of waveforms, it concludes that compared with orthogonal frequency division waveform, the crosscorrelation of orthogonal code waveform badly weakens the performance of spaceborne MIMO radar in GMTI (Ground moving target indication). Thus, by adopting orthogonal frequency division waveform, the basic principle of space-time-frequency adaptive processing is studied. Simulation results demonstrate the superiority of frequency division orthogonal MIMO radar in improving clutter suppression and GMTI performance.

  14. 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.

  15. Special paraunitary matrices, Cayley transform, and multidimensional orthogonal filter banks.

    PubMed

    Zhou, Jianping; Do, Minh N; Kovaĉević, Jelena

    2006-02-01

    We characterize and design multidimensional (MD) orthogonal filter banks using special paraunitary matrices and the Cayley transform. Orthogonal filter banks are represented by paraunitary matrices in the polyphase domain. We define special paraunitary matrices as paraunitary matrices with unit determinant. We show that every paraunitary matrix can be characterized by a special paraunitary matrix and a phase factor. Therefore, the design of paraunitary matrices (and thus of orthogonal filter banks) becomes the design of special paraunitary matrices, which requires a smaller set of nonlinear equations. Moreover, we provide a complete characterization of special paraunitary matrices in the Cayley domain, which converts nonlinear constraints into linear constraints. Our method greatly simplifies the design of MD orthogonal filter banks and leads to complete characterizations of such filter banks. PMID:16479821

  16. Direct Orthogonal Distance to Quadratic Surfaces in 3D.

    PubMed

    Lott, Gus K

    2014-09-01

    Discovering the orthogonal distance to a quadratic surface is a classic geometric task in vision, modeling, and robotics. I describe a simple, efficient, and stable direct solution for the orthogonal distance (foot-point) to an arbitrary quadratic surface from a general finite 3D point. The problem is expressed as the intersection of three quadratic surfaces, two of which are derived from the requirement of orthogonality of two non-coincident planes with the tangent plane to the quadric. A sixth order single-variable polynomial is directly generated in one coordinate of the surface point. The method detects intersection points at infinity and operates smoothly across all real quadratic surface classes. The method also geometrically detects continuums of orthogonal points (i.e., from the exact center of a sphere). I discuss algorithm performance, compare it to a state-of-the-art estimator, demonstrate the algorithm on synthetic data, and describe extension to arbitrary dimension. PMID:26352239

  17. 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

  18. A proof of van Aubel's theorem using orthogonal vectors

    NASA Astrophysics Data System (ADS)

    Glaister, P.

    2016-04-01

    We show how two linearly independent vectors can be used to construct two orthogonal vectors of equal magnitude in a simple way. The proof that the constructed vectors are orthogonal and of equal magnitude is a good exercise for students studying properties of scalar and vector triple products. We then show how this result can be used to prove van Aubel's theorem that relates the two line segments joining the centres of squares on opposite sides of a plane quadrilateral.

  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. 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.

  1. Orthogonal metals: The simplest non-Fermi liquids

    NASA Astrophysics Data System (ADS)

    Nandkishore, Rahul; Metlitski, Max A.; Senthil, T.

    2012-07-01

    We present a fractionalized metallic phase which is indistinguishable from the Fermi liquid in conductivity and thermodynamics, but is sharply distinct in one-electron properties, such as the electron spectral function. We dub this phase the “orthogonal metal.” The orthogonal metal and the transition to it from the Fermi liquid are naturally described using a slave-particle representation wherein the electron is expressed as a product of a fermion and a slave Ising spin. We emphasize that when the slave spins are disordered, the result is not a Mott insulator (as erroneously assumed in the prior literature), but rather the orthogonal metal. We construct prototypical ground-state wave functions for the orthogonal metal by modifying the Jastrow factor of Slater-Jastrow wave functions that describe ordinary Fermi liquids. We further demonstrate that the transition from the Fermi liquid to the orthogonal metal can, in some circumstances, provide a simple example of a continuous destruction of a Fermi surface with a critical Fermi surface appearing right at the critical point. We present exactly soluble models that realize an orthogonal metal phase, and the phase transition to the Fermi liquid. These models thus provide valuable solvable examples for phase transitions associated with the death of a Fermi surface.

  2. Expeditious oligosaccharide synthesis via selective, semi-orthogonal, and orthogonal activation

    PubMed Central

    Kaeothip, Sophon; Demchenko, Alexei V.

    2011-01-01

    Traditional strategies for oligosaccharide synthesis often require extensive protecting and/or leaving group manipulations between each glycosylation step, thereby increasing the total number of synthetic steps while decreasing the efficiency of the synthesis. In contrast, expeditious strategies allow for the rapid chemical synthesis of complex carbohydrates by minimizing extraneous chemical manipulations. Oligosaccharide synthesis by selective activation of one leaving group over another is one such expeditious strategy. Herein, the significant improvements that have recently emerged in the area of the selective activation are discussed. The development of orthogonal strategy further expands the scope of the selective activation methodology. Surveyed in this article, are representative examples wherein these excellent innovations have been applied to the synthesis of various oligosaccharide sequences. PMID:21663897

  3. 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

  4. 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.

  5. 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.

  6. Differential expression of argininosuccinate synthetase in serous and non‐serous ovarian carcinomas

    PubMed Central

    Cheon, Dong‐Joo; Walts, Ann E; Beach, Jessica A; Lester, Jenny; Bomalaski, John S; Walsh, Christine S; Ruprecht Wiedemeyer, W; Karlan, Beth Y

    2014-01-01

    Abstract The current standard of care for epithelial ovarian cancer does not discriminate between different histologic subtypes (serous, clear cell, endometrioid and mucinous) despite the knowledge that ovarian carcinoma subtypes do not respond uniformly to conventional platinum/taxane‐based chemotherapy. Exploiting addictions and vulnerabilities in cancers with distinguishable molecular features presents an opportunity to develop individualized therapies that may be more effective than the current ‘one size fits all' approach. One such opportunity is arginine depletion therapy with pegylated arginine deiminase, which has shown promise in several cancer types that exhibit low levels of argininosuccinate synthetase including hepatocellular and prostate carcinoma and melanoma. Based on the high levels of argininosuccinate synthetase previously observed in ovarian cancers, these tumours have been considered unlikely candidates for arginine depletion therapy. However, argininosuccinate synthetase levels have not been evaluated in the individual histologic subtypes of ovarian carcinoma. The current study is the first to examine the expression of argininosuccinate synthetase at the mRNA and protein levels in large cohorts of primary and recurrent ovarian carcinomas and ovarian cancer cell lines. We show that the normal fallopian tube fimbria and the majority of primary high‐grade and low‐grade serous ovarian carcinomas express high levels of argininosuccinate synthetase, which tend to further increase in recurrent tumours. In contrast to the serous subtype, non‐serous ovarian carcinoma subtypes (clear cell, endometrioid and mucinous) frequently lack detectable argininosuccinate synthetase expression. The in vitro sensitivity of ovarian cancer cell lines to arginine depletion with pegylated arginine deiminase was inversely correlated with argininosuccinate synthetase expression. Our data suggest that the majority of serous ovarian carcinomas are not susceptible

  7. 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. PMID:25801336

  8. Glutamine synthetase predicts adjuvant TACE response in hepatocellular carcinoma

    PubMed Central

    Zhang, Bo; Liu, Kai; Zhang, Jian; Dong, Liwei; Jin, Zhichao; Zhang, Xinji; Xue, Feng; He, Jia

    2015-01-01

    Background: Adjuvant transcatheter arterial chemoembolization (TACE) is associated with better outcome and reduced tumor recurrence in hepatocellular carcinoma (HCC) patients. This study aimed to investigate the relationship between glutamine synthetase (GS) expression and survival of HCC patients after postoperative adjuvant TACE. Methods: We retrospectively analyzed 554 HCC patients in two independent cohorts who underwent curative resection. Immunohistochemistry assay was used to investigate the expression of GS protein and evaluate the association with survival and the response to adjuvant TACE. Results: In training cohort, patients with low GS expression who received postoperative adjuvant TACE showed a better overall survival (OS) (P<0.001) and less early phase recurrence (P=0.016). Adjuvant TACE was an independent prognostic factor for 5-year OS (HR=0.408, 95% CI 0.261-0.639, P<0.001) and early phase recurrence (HR=0.592, 95% CI 0.376-0.931, P=0.023). The same result was confirmed in validation cohort. Patients with high GS expression in both cohorts did not have a significant response to adjuvant TACE in OS and early phase recurrence. Conclusions: GS status in tumor might be a useful tool in the selection of HCC patients who would be likely to benefit from postoperative adjuvant TACE. PMID:26884995

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

    PubMed Central

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

    2015-01-01

    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. DOI: http://dx.doi.org/10.7554/eLife.07607.001 PMID:26226635

  10. 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

  11. Isolation and characterization of glutamine synthetase genes in Chlamydomonas reinhardtii.

    PubMed

    Chen, Q; Silflow, C D

    1996-11-01

    To elucidate the role of glutamine synthetase (GS) in nitrogen assimilation in the green alga Chlamydomonas reinhardtii we used maize GS1 (the cytosolic form) and GS2 (the chloroplastic form) cDNAs as hybridization probes to isolate C. reinhardtii cDNA clones. The amino acid sequences derived from the C. reinhardtii clones have extensive homology with GS enzymes from higher plants. A putative amino-terminal transit peptide encoded by the GS2 cDNA suggests that the protein localizes to the chloroplast. Genomic DNA blot analysis indicated that GS1 is encoded by a single gene, whereas two genomic fragments hybridized to the GS2 cDNA probe. All GS2 cDNA clones corresponded to only one of the two GS2 genomic sequences. We provide evidence that ammonium, nitrate, and light regulate GS transcript accumulation in green algae. Our results indicate that the level of GS1 transcripts is repressed by ammonium but induced by nitrate. The level of GS2 transcripts is not affected by ammonium or nitrate. Expression of both GS1 and GS2 genes is regulated by light, but perhaps through different mechanisms. Unlike in higher plants, no decreased level of GS2 transcripts was detected when cells were grown under conditions that repress photorespiration. Analysis of GS transcript levels in mutants with defects in the nitrate assimilation pathway show that nitrate assimilation and ammonium assimilation are regulated independently. PMID:8938407

  12. The aminoacyl-tRNA synthetases of Drosophila melanogaster.

    PubMed

    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

  13. 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.

  14. Evidence for allosteric regulation of succinyl-CoA synthetase.

    PubMed Central

    Um, H D; Klein, C

    1993-01-01

    We have previously reported that distinctly different concentrations of GDP stimulate the phosphorylation and dephosphorylation of p36, the alpha-subunit of succinyl-CoA synthetase (SCS) in Dictyostelium discoideum. In this present study, we have investigated the mechanism underlying these dual effects of GDP. Dephosphorylation of p36 is induced by relatively high levels of GDP and is coincident with the formation of GTP. This indicates that, at high concentrations, GDP serves as a substrate of SCS. However, 100-fold lower concentrations of GDP, which do not bind to the catalytic site to induce SCS dephosphorylation, stimulate p36 phosphorylation. This stimulation is not diminished by dilution of the sample, and is retained during purification of the protein. Gel-filtration analyses indicate that SCS in our system behaves as a non-interacting alpha beta dimer, the hydrodynamic behaviour of which is not altered by the presence of added GDP. The data indicate that altered protein-protein interactions do not account for the stimulation of p36 phosphorylation by low GDP concentrations. We propose that GDP functions as an allosteric regulator of SCS, and experiments using guanosine 5'-[beta-thio]diphosphate (GDP[S]) are shown to distinguish further the allosteric and catalytic binding sites. Images Figure 1 Figure 2 Figure 3 Figure 5 PMID:8240297

  15. Evidence for allosteric regulation of succinyl-CoA synthetase.

    PubMed

    Um, H D; Klein, C

    1993-11-01

    We have previously reported that distinctly different concentrations of GDP stimulate the phosphorylation and dephosphorylation of p36, the alpha-subunit of succinyl-CoA synthetase (SCS) in Dictyostelium discoideum. In this present study, we have investigated the mechanism underlying these dual effects of GDP. Dephosphorylation of p36 is induced by relatively high levels of GDP and is coincident with the formation of GTP. This indicates that, at high concentrations, GDP serves as a substrate of SCS. However, 100-fold lower concentrations of GDP, which do not bind to the catalytic site to induce SCS dephosphorylation, stimulate p36 phosphorylation. This stimulation is not diminished by dilution of the sample, and is retained during purification of the protein. Gel-filtration analyses indicate that SCS in our system behaves as a non-interacting alpha beta dimer, the hydrodynamic behaviour of which is not altered by the presence of added GDP. The data indicate that altered protein-protein interactions do not account for the stimulation of p36 phosphorylation by low GDP concentrations. We propose that GDP functions as an allosteric regulator of SCS, and experiments using guanosine 5'-[beta-thio]diphosphate (GDP[S]) are shown to distinguish further the allosteric and catalytic binding sites. PMID:8240297

  16. Cloning of the glutamine synthetase gene from group B streptococci.

    PubMed

    Suvorov, A N; Flores, A E; Ferrieri, P

    1997-01-01

    The glnA gene from the human pathogen Streptococcus agalactiae was cloned from a genomic library prepared with the lambda phage vector lambdaDASHII. A 4.6-kb DNA fragment of one of the recombinant phages was subcloned in pUC18. This Escherichia coli clone expressed a 52-kDa protein encoded by a 1,341-bp open reading frame. The nucleotide sequence of the open reading frame and the deduced amino acid sequence shared a significant degree of homology with the sequences of other glutamine synthetases (GS). The highest homology was between our deduced protein and GS of gram-positive bacteria such as Bacillus subtilis, Bacillus cereus, and Staphylococcus aureus. Plasmids with the cloned streptococcal glnA were able to complement E. coli glnA mutants grown on minimal media. Rabbit antisera to streptococcal GS recombinant protein recognized not only the recombinant protein but also a similar-sized band in mutanolysin extracts of all group B streptococcal strains tested, regardless of polysaccharide type or surface protein profile. The amino acid sequence of the deduced protein had similarities to other streptococcal cell-surface-bound proteins. The possible functional role of the immunological features of streptococcal GS is discussed. PMID:8975911

  17. Adenine nucleotides as allosteric effectors of PEA seed glutamine synthetase

    SciTech Connect

    Unkefer, P.J.; Knight, T.J.

    1986-05-01

    The energy charge in the plant cell has been proposed as a regulator of glutamine synthetase (GS) activity. The authors have shown that 2.1 moles of ..gamma..(/sup 32/P)-ATP were bound/mole subunits of purified pea seed GS during complete inactivation with methionine sulfoximine. Since GS has one active site per subunit, the second binding site provides the potential for allosteric regulation of GS by adenine nucleotides. The authors have investigated the inhibition of the ATP-dependent synthetic activity by ADP and AMP. ADP and AMP cannot completely inhibit GS; but ATP does overcome the inhibition by ADP and AMP as shown by plots of % inhibition vs inhibitor concentration. This indicates that inhibition of GS by ADP or AMP is not completely due to competitive inhibition. In the absence of ADP or AMP, double reciprocal plots for ATP are linear below 10 mM; however, in the presence of either ADP or AMP these pots are curvilinear downwards. The ratio of Vm/asymptote is less than 1. The Hill number for ATP in the absence of ADP or AMP is 0.93 but decreases with increasing ADP or AMP to a value of 0.28 with 10 mM ADP. These data are consistent with negative cooperativity by ADP and AMP. Thus, as the ADP/ATP or AMP/ATP ratios are increased GS activity decreases. This is consistent with regulation of GS activity by energy charge in planta.

  18. 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. PMID:25896882

  19. 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

  20. Proteomic identification of glutamine synthetase as a differential marker for oligodendrogliomas and astrocytomas

    PubMed Central

    Zhuang, Zhengping; Qi, Meng; Li, Jie; Okamoto, Hiroaki; Xu, David S.; Iyer, Rajiv R.; Lu, Jie; Yang, Chunzhang; Weil, Robert J.; Vortmeyer, Alexander; Lonser, Russell R.

    2016-01-01

    Object Astrocytomas and oligodendrogliomas are primary CNS tumors that remain a challenge to differentiate histologically because of their morphological variability and because there is a lack of reliable differential diagnostic markers. To identify proteins that are differentially expressed between astrocytomas and oligodendrogliomas, the authors analyzed the proteomic expression patterns and identified uniquely expressed proteins in these neoplasms. Methods Proteomes of astrocytomas and oligodendrogliomas were analyzed using 2D gel electrophoresis and subsequent computerized gel analysis to detect differentially expressed proteins. The proteins were identified using high-performance liquid chromatography accompanied by tandem mass spectrometry. To determine the role of the differentially expressed proteins in astrocytes, undifferentiated glial cell cultures were treated with dibutyryl–cyclic adenosine monophosphate (cAMP). Results Two-dimensional gel electrophoresis revealed that glutamine synthetase was differentially expressed in astrocytomas and oligodendrogliomas. Western blot and immunohistochemical analyses confirmed the increased expression of glutamine synthetase in astrocytomas compared with oligodendrogliomas. Whereas glutamine synthetase expression was demonstrated across all grades of astrocytomas (Grade II–IV [15 tumors]) and oligoastrocytomas (4 tumors), it was expressed in only 1 oligodendroglioma (6% [16 tumors]). Treatment of undifferentiated glial cell cultures with dibutyryl-cAMP resulted in astrocyte differentiation that was associated with increased levels of glial fibrillary acidic protein and glutamine synthetase. Conclusions These data indicate that glutamine synthetase expression can be used to distinguish astrocytic from oligodendroglial tumors and may play a role in the pathogenesis of astrocytomas. PMID:21682567

  1. 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. PMID:27294598

  2. Isolation of the thymidylate synthetase gene (TMP1) by complementation in Saccharomyces cerevisiae

    SciTech Connect

    Taylor, G.R.; Barclay, B.J.; Storms, R.K.; Friesen, J.D.; Haynes, R.H.

    1982-04-01

    The structural gene (TMP1) for yeast thymidylate synthetase (thymidylate synthase; EC 2.1.1.45) was isolated from a chimeric plasmid bank by genetic complementation in Saccharomyces cerevisiae. Retransformation of the dTMP auxotroph GY712 and a temperature-sensitive mutant (cdc21) with purified plasmid (pTL1) yielded Tmp/sup +/ transformants at high frequency. In addition, the plasmid was tested for the ability to complement a bacterial thyA mutant that lacks functional thymidylate synthetase. Although it was not possible to select Thy/sup +/ transformants directly, it was found that all pTL1 transformants were phenotypically Thy/sup +/ after several generations of growth in nonselective conditions. Thus, yeast thymidylate synthetase is biologically active in Escherichia coli. Thymidylate synthetase was assayed in yeast cell lysates by high-pressure liquid chromatography to monitor the conversion of (6-/sup 3/H)dUMP to (6-/sup 3/H)dTMP. In protein extracts from the thymidylate auxotroph (tmpl-6) enzymatic conversion of dUMP to dTMP was barely detectable. Lysates of pTL1 transformants of this strain, however, had thymidylate synthetase activity that was comparable to that of the wild-type strain.

  3. A component of the multisynthetase complex is a multifunctional aminoacyl-tRNA synthetase.

    PubMed Central

    Cerini, C; Kerjan, P; Astier, M; Gratecos, D; Mirande, M; Sémériva, M

    1991-01-01

    In higher eukaryotes, nine aminoacyl-tRNA synthetases are associated within a multienzyme complex which is composed of 11 polypeptides with molecular masses ranging from 18 to 150 kDa. We have cloned and sequenced a cDNA from Drosophila encoding the largest polypeptide of this complex. We demonstrate here that the corresponding protein is a multifunctional aminoacyl-tRNA synthetase. It is composed of three major domains, two of them specifying distinct synthetase activities. The amino and carboxy-terminal domains were expressed separately in Escherichia coli, and were found to catalyse the aminoacylation of glutamic acid and proline tRNA species, respectively. The central domain is made of six 46 amino acid repeats. In prokaryotes, these two aminoacyl-tRNA synthetases are encoded by distinct genes. The emergence of a multifunctional synthetase by a gene fusion event seems to be a specific, but general attribute of all higher eukaryotic cells. This type of structural organization, in relation to the occurrence of multisynthetase complexes, could be a mechanism to integrate several catalytic domains within the same particle. The involvement of the internal repeats in mediating complex assembly is discussed. Images PMID:1756734

  4. Identification and Functional Characterization of a Novel Bacterial Type Asparagine Synthetase A

    PubMed Central

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

    2014-01-01

    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. PMID:24610810

  5. 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

  6. Membrane Anchoring of Aminoacyl-tRNA Synthetases by Convergent Acquisition of a Novel Protein Domain*

    PubMed Central

    Olmedo-Verd, Elvira; Santamaría-Gómez, Javier; Ochoa de Alda, Jesús A. G.; Ribas de Pouplana, Lluis; Luque, Ignacio

    2011-01-01

    Four distinct aminoacyl-tRNA synthetases (aaRSs) found in some cyanobacterial species contain a novel protein domain that bears two putative transmembrane helices. This CAAD domain is present in glutamyl-, isoleucyl-, leucyl-, and valyl-tRNA synthetases, the latter of which has probably recruited the domain more than once during evolution. Deleting the CAAD domain from the valyl-tRNA synthetase of Anabaena sp. PCC 7120 did not significantly modify the catalytic properties of this enzyme, suggesting that it does not participate in its canonical tRNA-charging function. Multiple lines of evidence suggest that the function of the CAAD domain is structural, mediating the membrane anchorage of the enzyme, although membrane localization of aaRSs has not previously been described in any living organism. Synthetases containing the CAAD domain were localized in the intracytoplasmic thylakoid membranes of cyanobacteria and were largely absent from the plasma membrane. The CAAD domain was necessary and apparently sufficient for protein targeting to membranes. Moreover, localization of aaRSs in thylakoids was important under nitrogen limiting conditions. In Anabaena, a multicellular filamentous cyanobacterium often used as a model for prokaryotic cell differentiation, valyl-tRNA synthetase underwent subcellular relocation at the cell poles during heterocyst differentiation, a process also dependent on the CAAD domain. PMID:21965654

  7. 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.

  8. Structural plasticity of an aminoacyl-tRNA synthetase active site

    PubMed Central

    Turner, James M.; Graziano, James; Spraggon, Glen; Schultz, Peter G.

    2006-01-01

    Recently, tRNA aminoacyl-tRNA synthetase pairs have been evolved that allow one to genetically encode a large array of unnatural amino acids in both prokaryotic and eukaryotic organisms. We have determined the crystal structures of two substrate-bound Methanococcus jannaschii tyrosyl aminoacyl-tRNA synthetases that charge the unnatural amino acids p-bromophenylalanine and 3-(2-naphthyl)alanine (NpAla). A comparison of these structures with the substrate-bound WT synthetase, as well as a mutant synthetase that charges p-acetylphenylalanine, shows that altered specificity is due to both side-chain and backbone rearrangements within the active site that modify hydrogen bonds and packing interactions with substrate, as well as disrupt the α8-helix, which spans the WT active site. The high degree of structural plasticity that is observed in these aminoacyl-tRNA synthetases is rarely found in other mutant enzymes with altered specificities and provides an explanation for the surprising adaptability of the genetic code to novel amino acids. PMID:16618920

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

    NASA Astrophysics Data System (ADS)

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

    2016-08-01

    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.

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

    PubMed

    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

  11. 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.

  12. 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

    2016-08-01

    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. PMID:27434548

  13. Orthogonal chemical functionalization of patterned gold on silica surfaces

    PubMed Central

    Léonard, Didier; Le Mogne, Thierry; Zuttion, Francesca; Chevalier, Céline; Phaner-Goutorbe, Magali; Souteyrand, Éliane

    2015-01-01

    Summary Single-step orthogonal chemical functionalization procedures have been developed with patterned gold on silica surfaces. Different combinations of a silane and a thiol were simultaneously deposited on a gold/silica heterogeneous substrate. The orthogonality of the functionalization (i.e., selective grafting of the thiol on the gold areas and the silane on the silica) was demonstrated by X-ray photoelectron spectroscopy (XPS) as well as time-of-flight secondary ion mass spectrometry (ToF–SIMS) mapping. The orthogonal functionalization was used to immobilize proteins onto gold nanostructures on a silica substrate, as demonstrated by atomic force microscopy (AFM). These results are especially promising in the development of future biosensors where the selective anchoring of target molecules onto nanostructured transducers (e.g., nanoplasmonic biosensors) is a major challenge. PMID:26734519

  14. Orthogonal chemical functionalization of patterned gold on silica surfaces.

    PubMed

    Palazon, Francisco; Léonard, Didier; Le Mogne, Thierry; Zuttion, Francesca; Chevalier, Céline; Phaner-Goutorbe, Magali; Souteyrand, Éliane; Chevolot, Yann; Cloarec, Jean-Pierre

    2015-01-01

    Single-step orthogonal chemical functionalization procedures have been developed with patterned gold on silica surfaces. Different combinations of a silane and a thiol were simultaneously deposited on a gold/silica heterogeneous substrate. The orthogonality of the functionalization (i.e., selective grafting of the thiol on the gold areas and the silane on the silica) was demonstrated by X-ray photoelectron spectroscopy (XPS) as well as time-of-flight secondary ion mass spectrometry (ToF-SIMS) mapping. The orthogonal functionalization was used to immobilize proteins onto gold nanostructures on a silica substrate, as demonstrated by atomic force microscopy (AFM). These results are especially promising in the development of future biosensors where the selective anchoring of target molecules onto nanostructured transducers (e.g., nanoplasmonic biosensors) is a major challenge. PMID:26734519

  15. Orthogonal test and experimental study on fire floating pump

    NASA Astrophysics Data System (ADS)

    Liu, J. R.; Zheng, J. F.; Fu, D. P.; Wang, P.

    2013-12-01

    In order to develop high efficiency fire floating pump, 250YYB-250 fire floating pump was taken as an example. The orthogonal experiment of L9 (34), which contains factors with three levels of blade numbers of impeller, outlet angle, impeller fold-angle, was performed to design nine types of impellers. Numerical simulation of whole flow field based on Fluent was adopted to perform an orthogonal test, the order of geometric parameters affects the performance of fire floating pump with complex impeller. The best design scheme for pump model was acquired. Meanwhile, the optimized design scheme was determined, and corresponding test was carried out. It demonstrated that the efficiency of the final optimal design model pump at rated flow point is of 85%. The efficiency is higher than the national standards, which verified the feasibility of the method of orthogonal design in pump design.

  16. Scattering equations and Kawai-Lewellen-Tye orthogonality

    NASA Astrophysics Data System (ADS)

    Cachazo, Freddy; He, Song; Yuan, Ellis Ye

    2014-09-01

    Several recent developments point to the fact that rational maps from n-punctured spheres to the null cone of D-dimensional momentum space provide a natural language for describing the scattering of massless particles in D dimensions. In this paper we identify and study equations relating the kinematic invariants sab and the puncture locations σc, which we call the scattering equations. We provide an inductive algorithm in the number of particles for their solutions and prove a remarkable property which we call Kawai-Lewellen-Tye (KLT) orthogonality. In a nutshell, KLT orthogonality means that "Parke-Taylor" vectors constructed from the solutions to the scattering equations are mutually orthogonal with respect to the KLT bilinear form. We end with comments on possible connections to gauge theory and gravity amplitudes in any dimension and to the high-energy limit of string theory amplitudes.

  17. 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.

  18. Calculation on flux-MMF relationship of orthogonal-core

    SciTech Connect

    Tajima, K.; Kaga, A.; Anazawa, Y. ); Ichinokura, O. )

    1993-03-01

    Orthogonal-cores have various potential applications, for instance in parametric transformers and dc-ac converters. The operating characteristics of the devices can be calculated on the basis of the measured relationship of flux to MMF of the orthogonal-core. To achieve optimal design of the applied device, the relationship of flux to MMF must be determined; however, this involves solving a three dimensional nonlinear problem. In this paper, the authors calculate the flux-MMF relationship based on a magnetic circuit model for the orthogonal-core. The computed results agree well with experiment. The method of this study is shown to be valid for calculation of characteristics and useful for optimal design of application devices.

  19. 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.

  20. 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.

  1. 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

  2. Measurement matrix optimization method based on matrix orthogonal similarity transformation

    NASA Astrophysics Data System (ADS)

    Pan, Jinfeng

    2016-05-01

    Optimization of the measurement matrix is one of the important research aspects of compressive sensing theory. A measurement matrix optimization method is presented based on the orthogonal similarity transformation of the information operator's Gram matrix. In terms of the fact that the information operator's Gram matrix is a singular symmetric matrix, a simplified orthogonal similarity transformation is deduced, and thus the simplified diagonal matrix that is orthogonally similar to it is obtained. Then an approximation of the Gram matrix is obtained by letting all the nonzero diagonal entries of the simplified diagonal matrix equal their average value. Thus an optimized measurement matrix can be acquired according to its relationship with the information operator. Results of experiments show that the optimized measurement matrix compared to the random measurement matrix is less coherent with dictionaries. The relative signal recovery error also declines when the proposed measurement matrix is utilized.

  3. Further characterization of Escherichia coli alanyl-tRNA synthetase.

    PubMed

    Sood, S M; Slattery, C W; Filley, S J; Wu, M X; Hill, K A

    1996-04-15

    Selected physical and thermodynamic parameters for Escherichia coli alanyl-tRNA synthetase (AlaRS) have been determined primarily to assess the quaternary structure of this enzyme. The extinction coefficient (epsilon) at 280 nm was determined experimentally to be 0.71 ml mg-1 cm-1, and the partial specific volume (nu) was calculated from the amino acid composition to be 0.73 ml g-1. From viscosity experiments the intrinsic viscosity (eta) of AlaRS was extrapolated to be 3.4 ml g-1 and the degree of hydration (delta 1) estimated to be 0.67 gH2O g(-1)(AlaRS). Laser light-scattering studies indicated some heterogeneity; a radius of 6.3 nm was calculated for the major fraction with a diffusion coefficient (D20,W) of 3.89 x 10(-7) cm2 s-1. In 50 mM Hepes, pH 7.5, 20 mM KCl, 2 mM 2-mercaptoethanol and at a protein concentration of 4.2 mg ml-1 the sedimentation coefficient (S20,W) was 6.36 S; this value increased slightly when the protein concentration was decreased. The combination of S20,W and D20,W under these conditions yielded a molecular weight of approximately 186,000 Da, corresponding to a dimer. The S20,W was virtually independent of temperature in the range of 10-37 degrees C, while an Arrhenius plot of aminoacylation activity was biphasic. The isoelectric point was determined experimentally to be 4.9. Sedimentation equilibrium data were best fit to a decamer association complex in which dimeric AlaRS is the predominant species at 25 degrees C. PMID:8645007

  4. Antimalarial Benzoxaboroles Target Plasmodium falciparum Leucyl-tRNA Synthetase.

    PubMed

    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; Rosenthal, Philip J

    2016-08-01

    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 [(14)C]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

  5. Role of thymidylate synthetase activity in development of methotrexate cytotoxicity.

    PubMed Central

    Moran, R G; Mulkins, M; Heidelberger, C

    1979-01-01

    Methotrexate (MTX) inhibition of the growth of mouse or human leukemia cells in culture was partially prevented by either thymidine (dThd) or hypoxanthine. 5-Fluoro-2'-deoxyuridine (FdUrd) also decreased the growth-inhibitory potency of MTX in the presence of small concentrations of 5-formyltetrahydrofolate (citrovorum factor) and sufficient exogenous dThd to support the synthesis of thymidylate nucleotides by salvage mechanisms. In addition, citrovorum factor-induced reversal of MTX was several orders of magnitude more efficient in the presence of both FdUrd and dThd than in the presence of dThd alone or in the absence of both nucleosides. Likewise, the presence of FdUrd (3 microM) and dThd (5.6 microM) completely prevented the lethality of 0.3 mM MTX to L1210 cells in culture medium supplemented with micromolar concentrations of citrovorum factor. We propose that this protection against the cytotoxic effects of MTX by dThd, hypoxanthine, and FdUrd have a common biochemical mechanism--namely, inhibition of the de novo synthesis of thymidylate by either a direct [FdUrd; inhibition of thymidylate synthetase (thymidylate synthase; 5,10-methylenetetrahydrofolate:dUMP C-methyl-transferase, EC 2.1.1.45)] or indirect (dThd and hypoxanthine; feedback inhibition by anabolites on ribonucleotide reductase and deoxycytidylate deaminase) effect. The resultant decreased rate of loss of reduced folates due to de novo thymidylate synthesis would allow a higher degree of inhibition of dihydrofolate reductase to be endured without damage to the cell. PMID:160558

  6. Pathogenic implications of human mitochondrial aminoacyl-tRNA synthetases.

    PubMed

    Schwenzer, Hagen; Zoll, Joffrey; Florentz, Catherine; Sissler, Marie

    2014-01-01

    Mitochondria are considered as the powerhouse of eukaryotic cells. They host several central metabolic processes fueling the oxidative phosphorylation pathway (OXPHOS) that produces ATP from its precursors ADP and inorganic phosphate Pi (PPi). The respiratory chain complexes responsible for the OXPHOS pathway are formed from complementary sets of protein subunits encoded by the nuclear genome and the mitochondrial genome, respectively. The expression of the mitochondrial genome requires a specific and fully active translation machinery from which aminoacyl-tRNA synthetases (aaRSs) are key actors. Whilst the macromolecules involved in mammalian mitochondrial translation have been under investigation for many years, there has been an explosion of interest in human mitochondrial aaRSs (mt-aaRSs) since the discovery of a large (and growing) number of mutations in these genes that are linked to a variety of neurodegenerative disorders. Herein we will review the present knowledge on mt-aaRSs in terms of their biogenesis, their connection to mitochondrial respiration, i.e., the respiratory chain (RC) complexes, and to the mitochondrial translation machinery. The pathology-related mutations detected so far are described, with special attention given to their impact on mt-aaRSs biogenesis, functioning, and/or subsequent activities. The collected data to date shed light on the diverse routes that are linking primary molecular possible impact of a mutation to its phenotypic expression. It is envisioned that a variety of mechanisms, inside and outside the translation machinery, would play a role on the heterogeneous manifestations of mitochondrial disorders. PMID:23824528

  7. Proteasomal degradation of glutamine synthetase regulates schwann cell differentiation.

    PubMed

    Saitoh, Fuminori; Araki, Toshiyuki

    2010-01-27

    Rapid saltatory nerve conduction is facilitated by myelin structure, which is composed of Schwann cells in the peripheral nervous system. Schwann cells drastically change their phenotype following peripheral nerve injury. These phenotypic changes are required for efficient degeneration/regeneration. We previously identified ZNRF1 as an E3 ubiquitin ligase containing a RING finger motif, whose expression is upregulated in the Schwann cells following nerve injury. This suggested that posttranscriptional regulation of protein expression in Schwann cells may be involved in their phenotypic changes during nerve degeneration/regeneration. Here we report the identification of glutamine synthetase (GS), an enzyme that synthesizes glutamine using glutamate and ammonia, as a substrate for E3 activity of ZNRF1 in Schwann cells. GS is known to be highly expressed in differentiated Schwann cells, but its functional significance has remained unclear. We found that during nerve degeneration/regeneration, GS expression is controlled mostly by ZNRF1-dependent proteasomal degradation. We also found that Schwann cells increase oxidative stress upon initiation of nerve degeneration, which promotes carbonylation and subsequent degradation of GS. Surprisingly, we discovered that GS expression regulates Schwann cell differentiation; i.e., increased GS expression promotes myelination via its enzymatic activity. Among the substrates and products of GS, increased glutamate concentration inhibited myelination and yet promoted Schwann cell proliferation by activating metabotropic glutamate receptor signaling. This would suggest that GS may exert its effect on Schwann cell differentiation by regulating glutamate concentration. These results indicate that the ZNRF1-GS system may play an important role in correlating Schwann cell metabolism with its differentiation. PMID:20107048

  8. 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.

  9. Universal Continuous-Variable State Orthogonalizer and Qubit Generator

    NASA Astrophysics Data System (ADS)

    Coelho, Antonio S.; Costanzo, Luca S.; Zavatta, Alessandro; Hughes, Catherine; Kim, M. S.; Bellini, Marco

    2016-03-01

    We experimentally demonstrate a universal strategy for producing a quantum state that is orthogonal to an arbitrary, infinite-dimensional, pure input one, even if only a limited amount of information about the latter is available. Arbitrary coherent superpositions of the two mutually orthogonal states are then produced by a simple change in the experimental parameters. We use input coherent states of light to illustrate two variations of the method. However, we show that the scheme works equally well for arbitrary input fields and constitutes a universal procedure, which may thus prove a useful building block for quantum state engineering and quantum information processing with continuous-variable qubits.

  10. Equipment and preliminary results for orthogonal pressurized planar electrochromatography.

    PubMed

    Dzido, Tadeusz H; Lopaciuk, Eryk; Płocharz, Paweł W; Chomicki, Adam; Zembrzycka, Magdalena; Frank, Hartmut

    2014-03-21

    We report combination of overpressured layer chromatography (OPLC) and pressurized planar electrochromatography (PPEC) techniques into a single technique in which both OPLC and PPEC processes proceed simultaneously and orthogonally. The separation process with this new technique is performed in adsorbent layer of a chromatographic plate, which is equipped with special sealing margin on its whole periphery and closed under pressure in special chamber. We have named this separation technique as orthogonal pressurized planar electrochromatography (OPPEC). Examples of analytical and micropreparative (continuous) OPPEC separations are demonstrated. PMID:24572544

  11. Nineteen-year follow-up of a patient with severe glutathione synthetase deficiency.

    PubMed

    Atwal, Paldeep S; Medina, Casey R; Burrage, Lindsay C; Sutton, V Reid

    2016-07-01

    Glutathione synthetase deficiency is a rare autosomal recessive disorder resulting in low levels of glutathione and an increased susceptibility to oxidative stress. Patients with glutathione synthetase deficiency typically present in the neonatal period with hemolytic anemia, metabolic acidosis and neurological impairment. Lifelong treatment with antioxidants has been recommended in an attempt to prevent morbidity and mortality associated with the disorder. Here, we present a 19-year-old female who was diagnosed with glutathione synthetase deficiency shortly after birth and who has been closely followed in our metabolic clinic. Despite an initial severe presentation, she has had normal intellectual development and few complications of her disorder with a treatment regimen that includes polycitra (citric acid, potassium citrate and sodium citrate), vitamin C, vitamin E and selenium. PMID:26984560

  12. Constitutive Expression of Enniatin Synthetase during Fermentative Growth of Fusarium scirpi

    PubMed Central

    Billich, Andreas; Zocher, Rainer

    1988-01-01

    The production of enniatins by Fusarium scirpi during fermentative growth in submerged cultures was measured. The fungus produced the antibiotic during mycelial growth, but not during the stationary phase of cultivation. By contrast, enniatin synthetase, the enzyme responsible for enniatin synthesis, was present during growth, during the stationary phase, and even in spores. Similarly, the enniatin synthetase mRNA was present at every stage of the cultivation of the fungus. Therefore, this multifunctional peptide synthetase is a constitutive enzyme, the expression of which is not regulated by any specific mechanism. The findings stand in contrast to the common assumption that production of secondary metabolites underlies regulatory control, leading to separation of the trophophase and the idiophase. Images PMID:16347758

  13. A Bacterial Ortholog of Class II Lysyl-tRNA Synthetase Activates Lysine

    PubMed Central

    Ambrogelly, Alexandre; O’Donoghue, Patrick; Söll, Dieter; Moses, Sharath

    2010-01-01

    Aminoacyl-tRNA synthetases produce aminoacyl-tRNAs, essential substrates for accurate protein synthesis. Beyond their central role in translation some of these enzymes or their orthologs are recruited for alternative functions, not always related to their primary cellular role. We investigate here the enzymatic properties of GenX (also called PoxA and YjeA), an ortholog of bacterial class II lysyl-tRNA synthetase. GenX is present in most Gram-negative bacteria and is homologous to the catalytic core of lysyl-tRNA synthetase, but it lacks the amino terminal anticodon binding domain of the latter enzyme. We show that, in agreement with its well-conserved lysine binding site, GenX can activate in vitro L-lysine and lysine analogs, but does not acylate tRNALys or other cellular RNAs. PMID:20580719

  14. Variations in the Localization of Acetyl-Coenzyme A Synthetase in Aerobic Yeast Cells

    PubMed Central

    Klein, Harold P.; Jahnke, Linda

    1971-01-01

    In cells of Saccharomyces cerevisiae growing aerobically for 24 hr, acetyl-coenzyme A synthetase [acetate: CoA ligase (AMP), EC 6.2.1.1] was localized principally in the microsomal fraction. On density gradients, the enzyme in such cells behaved as a low-density particle, readily separable from the soluble proteins. After 48 hr of incubation, the cells showed a bimodal distribution of enzyme, with most of the activity now sedimenting with the mitochondrial fraction and only a smaller amount with the microsomal fraction. By using density gradients, two forms of synthetase were obtained from these cells: one band denser and the other band less dense than the intact mitochondria. In all preparations containing synthetase activity, appreciable levels of phospholipids were also detected. Images PMID:4102333

  15. Alteration of the Bacillus subtilis glutamine synthetase results in overproduction of the enzyme.

    PubMed Central

    Dean, D R; Hoch, J A; Aronson, A I

    1977-01-01

    A mutational leading to glutamine auxotrophy was located near a 5-fluorouracil resistance marker in the citB-thyA region of the Bacillus subtilis chromosome. This mutation resulted in a glutamine synthetase with altered kinetic and feedback properties. The specific activity of manganese-stimulated glutamine synthetase activity in crude extracts was 18-fold higher, and the magnesium-stimulated activity was about 30% that of the wild type. Quantitation of the enzyme by precipitation with antibody prepared against pure enzyme confirmed the presence of high enzyme levels in the mutant. This mutation is very closely linked (recombination index of 0.03) to another glutamine auxotroph containing enzyme with altered electrophoretic and heat sensitivity properties. Mutations in the structural gene for glutamine synthetase may result not only in altered catalytic and regulatory properties but also in altered production of the enzyme. Images PMID:19424

  16. Isolation and characterisation of a ferrirhodin synthetase gene from the sugarcane pathogen Fusarium sacchari.

    PubMed

    Munawar, Asifa; Marshall, James W; Cox, Russell J; Bailey, Andy M; Lazarus, Colin M

    2013-02-11

    FSN1, a gene isolated from the sugar-cane pathogen Fusarium sacchari, encodes a 4707-residue nonribosomal peptide synthetase consisting of three complete adenylation, thiolation and condensation modules followed by two additional thiolation and condensation domain repeats. This structure is similar to that of ferricrocin synthetase, which makes a siderophore that is involved in intracellular iron storage in other filamentous fungi. Heterologous expression of FSN1 in Aspergillus oryzae resulted in the accumulation of a secreted metabolite that was identified as ferrirhodin. This siderophore was found to be present in both mycelium and culture filtrates of F. sacchari, whereas ferricrocin is found only in the mycelium, thus suggesting that ferricrocin is an intracellular storage siderophore in F. sacchari, whereas ferrirhodin is used for iron acquisition. To our knowledge, this is the first report to characterise a ferrirhodin synthetase gene functionally. PMID:23307607

  17. A human aminoacyl-tRNA synthetase as a regulator of angiogenesis.

    PubMed

    Wakasugi, Keisuke; Slike, Bonnie M; Hood, John; Otani, Atsushi; Ewalt, Karla L; Friedlander, Martin; Cheresh, David A; Schimmel, Paul

    2002-01-01

    Aminoacyl-tRNA synthetases catalyze the first step of protein synthesis. It was shown recently that human tyrosyl-tRNA synthetase (TyrRS) can be split into two fragments having distinct cytokine activities, thereby linking protein synthesis to cytokine signaling pathways. Tryptophanyl-tRNA synthetase (TrpRS) is a close homologue of TyrRS. A natural fragment, herein designated as mini TrpRS, was shown by others to be produced by alternative splicing. Production of this fragment is reported to be stimulated by IFN-gamma, a cytokine that also stimulates production of angiostatic factors. Mini TrpRS is shown here to be angiostatic in a mammalian cell culture system, the chicken embryo, and two independent angiogenesis assays in the mouse. The full-length enzyme is inactive in the same assays. Thus, protein synthesis may be linked to the regulation of angiogenesis by a natural fragment of TrpRS. PMID:11773626

  18. A human aminoacyl-tRNA synthetase as a regulator of angiogenesis

    PubMed Central

    Wakasugi, Keisuke; Slike, Bonnie M.; Hood, John; Otani, Atsushi; Ewalt, Karla L.; Friedlander, Martin; Cheresh, David A.; Schimmel, Paul

    2002-01-01

    Aminoacyl-tRNA synthetases catalyze the first step of protein synthesis. It was shown recently that human tyrosyl-tRNA synthetase (TyrRS) can be split into two fragments having distinct cytokine activities, thereby linking protein synthesis to cytokine signaling pathways. Tryptophanyl-tRNA synthetase (TrpRS) is a close homologue of TyrRS. A natural fragment, herein designated as mini TrpRS, was shown by others to be produced by alternative splicing. Production of this fragment is reported to be stimulated by IFN-γ, a cytokine that also stimulates production of angiostatic factors. Mini TrpRS is shown here to be angiostatic in a mammalian cell culture system, the chicken embryo, and two independent angiogenesis assays in the mouse. The full-length enzyme is inactive in the same assays. Thus, protein synthesis may be linked to the regulation of angiogenesis by a natural fragment of TrpRS. PMID:11773626

  19. Neurospora crassa glutamine synthetase. Translation of specific messenger ribonucleic acid in a cell-free system derived from rabbit reticulocytes.

    PubMed

    Palacios, R; Campomanes, M; Quinto, C

    1977-05-10

    The total reticulocyte lysate cell-free protein-synthesizing system was incubated in the presence of Neurospora crassa RNA. With the aid of an antibody directed against purified N. crassa glutamine synthetase, the synthesis of a specific protein was detected. This protein precipitates with antiglutamine synthetase using both direct and indirect procedures, migrates with the same molecular weight as the monomer of N. crassa glutamine synthetase when subjected to acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, and chromatographs as N. crassa glutamine synthetase on anthranilate-bound Sepharose. These data indicate the translation of the mRNA that codes for N. crassa glutamine synthetase. This RNA behaves as poly(A)-containing material when fractionated on oly(U)-Sepha-rose. PMID:16013

  20. Properties and substrate specificity of the leucyl-, the threonyl- and the valyl-transfer-ribonucleic acid synthetases from Aesculus species

    PubMed Central

    Anderson, J. W.; Fowden, L.

    1970-01-01

    1. Leucyl- and threonyl-tRNA synthetases were partially purified up to 100-fold and 30-fold respectively from cotyledons of Aesculus hippocastanum and were largely separated from the other aminoacyl-tRNA synthetases. Valyl-tRNA synthetase was purified 25-fold from cotyledons of Aesculus californica. 2. Some properties are reported for the three enzymes when assayed by the [32P]pyrophosphate-ATP exchange technique. 3. β-(Methylenecyclopropyl)alanine, isoleucine, azaleucine, norleucine and γ-hydroxynorvaline acted as alternative substrates for the leucyl-tRNA synthetase; the enzyme's affinity for β-(methylenecyclopropyl)-alanine and for isoleucine was about 80-fold less than that exhibited for leucine. 4. α-Cyclopropylglycine and α-cyclobutylglycine acted as alternative substrates for the valyl-tRNA synthetase. PMID:5493505

  1. Acyl-CoA synthetase catalyzes the synthesis of diadenosine hexaphosphate (Ap6A).

    PubMed

    Fontes, R; Günther Sillero, M A; Sillero, A

    1999-03-01

    The synthesis of diadenosine hexaphosphate (Ap6A), a potent vasoconstrictor, is catalyzed by acyl-CoA synthetase from Pseudomonas fragi. In a first step AMP is transferred from ATP to tetrapolyphosphate (P4) originating adenosine pentaphosphate (p5A) which, subsequently, is the acceptor of another AMP moiety from ATP generating diadenosine hexaphosphate (Ap6A). Diadenosine pentaphosphate (Ap5A) and diadenosine tetraphosphate (Ap4A) were also synthesized in the course of the reaction. In view of the variety of biological effects described for these compounds the potential capacity of synthesis of diadenosine polyphosphates by the mammalian acyl-CoA synthetases may be relevant. PMID:10385004

  2. Acute Onset Anti-Synthetase Syndrome With Pericardial Effusion and Non-Specific Interstitial Pneumonia

    PubMed Central

    Shah, Aditya; Patel, Samir R.

    2016-01-01

    Anti-synthetase syndrome (AS) is a clinical entity which is described classically by the triad of interstitial lung disease (ILD), inflammatory myositis and presence of aminoacyl-tRNA synthetase antibodies (ASA). We describe a rare presentation of this condition with regard to the uncharacteristically acute nature of presentation, acute decompensation in clinical condition, development of acute interstitial pneumonitis requiring rescue extracorporeal membrane oxygenation (ECMO) and accompaniment of significant pericardial effusion on presentation, followed by rapid improvement with initiation of steroids. PMID:27540445

  3. Mutants of Phycomyces blakesleeanus Defective in Acetyl-CoA Synthetase

    PubMed

    Garre; Torres-Martinez

    1996-03-01

    Nine mutants of the filamentous fungus Phycomyces blakesleeanus have been isolated on the basis of their resistance to fluoroacetate. None of the isolates uses acetate as the sole carbon source. Genetic complementation experiments revealed that all the mutants belong to the same complementation group. Biochemical analysis indicated that the acetate-induced acetyl-CoA synthetase activity is abolished in all nine mutants, thus suggesting that they are affected in the gene coding for acetyl-CoA synthetase (facA). PMID:8812287

  4. Isolation of the facA (acetyl-CoA synthetase) gene of Phycomyces blakesleeanus.

    PubMed

    Garre, V; Murillo, F J; Torres-Martínez, S

    1994-08-01

    A 5.6 kb DNA fragment from the fungus Phycomyces blakesleeanus has been cloned and sequenced. The fragment contains a gene that probably codes for the enzyme acetyl-coenzyme A synthetase (facA). The amino acid sequence deduced for the P. blakesleeanus protein is highly homologous to those of acetyl-coA-synthetases from other organisms. When placed under the control of a constitutive promoter from Aspergillus nidulans, the cloned gene complemented a facA- mutation of this organism. In P. blakesleeanus, the expression of facA is induced by acetate. PMID:7914670

  5. Acute Onset Anti-Synthetase Syndrome With Pericardial Effusion and Non-Specific Interstitial Pneumonia.

    PubMed

    Shah, Aditya; Patel, Samir R

    2016-09-01

    Anti-synthetase syndrome (AS) is a clinical entity which is described classically by the triad of interstitial lung disease (ILD), inflammatory myositis and presence of aminoacyl-tRNA synthetase antibodies (ASA). We describe a rare presentation of this condition with regard to the uncharacteristically acute nature of presentation, acute decompensation in clinical condition, development of acute interstitial pneumonitis requiring rescue extracorporeal membrane oxygenation (ECMO) and accompaniment of significant pericardial effusion on presentation, followed by rapid improvement with initiation of steroids. PMID:27540445

  6. Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. Binding of pyridoxal phosphate to 5-aminolaevulinate synthetase.

    PubMed Central

    Davies, R C; Neuberger, A

    1979-01-01

    1. Pyridoxal 5'-phosphate is a cofactor essential for the enzymic activity of aminolaevulinate synthetase from Rhodopseudomonas spheroides. It also aids activation of the low-activity enzyme by trisulphides such as cystine trisulphide, whereas inactivation of enzyme is facilitated by its absence. 2. The fluorescence spectrum of purified high-activity enzyme is that expected for a pyridoxal phosphate--Schiff base, but the firmly bound cofactor does not appear to be at the active centre. In dilute solutions of enzyme this grouping is inaccessible to nucleophiles such as glycine, hydroxylamine, borohydride and cyanide, at pH 7.4. 3. An active-centre Schiff base is formed between enzyne and added pyridoxal phosphate, which is accessible to nucleophiles. Concentrated solutions of this enzyme--Schiff base on treatment with glycine yield apo- and semi-apoenzyme, which can re-bind pyridoxal phosphate. 4. Two types of binding of pyridoxal phosphate are distinguishable in dilute solution of enzyme, but these become indistinguishable when concentrated solutions are treated with cofactor. A change occurs in the susceptibility towards borohydride of the fluorescence of the "structural" pyridoxal phosphate. 5. One or two molecules of cofactor are bound per subunit of mol. wt. 50 000 in semiapo- or holo-enzyme. The fluorescence of pyridoxamine phosphate covalently bound to enzyme also indicates one to two nmol of reducible Schiff base per 7000 units of activity in purified and partially purified samples of enzyme. 6. Cyanide does not convert high-activity into low-activity enzyme, but with the enzyme-pyridoxal phosphate complex it forms a yellow fluorescent derivative that is enzymically active. PMID:312102

  7. Post-transcriptional regulation of S-adenosylmethionine synthetase from its stored mRNA in germinated wheat embryos.

    PubMed

    Mathur, M; Saluja, D; Sachar, R C

    1991-06-24

    About 2-3-fold stimulation of S-adenosylmethionine synthetase was witnessed in germinated wheat embryos (48 h). The enhancement of enzyme activity was significantly inhibited by cycloheximide and amino acid analogues. Simultaneous addition of corresponding amino acids alleviated the inhibitory effect of amino acid analogues. Conclusive proof for the de novo synthesis of S-adenosylmethionine synthetase was obtained by labelling this enzyme with [35SO4]2- in vivo. Thus de novo enzyme synthesis seemed necessary for the rise in activity of AdoMet synthetase in wheat embryos. Curiously, blocking of transcription with cordycepin failed to repress the de novo synthesis of AdoMet synthetase in germinated wheat embryos. We envisage the presence of stored mRNA for AdoMet synthetase in wheat embryos. Thus the regulation of this enzyme occurs at the post-transcriptional level. L-Methionine, which is one of the substrates of AdoMet synthetase, stimulated the enzyme activity (2-2.4-fold) over that observed in control germinated embryos. L-Methionine promotes increased de novo synthesis of AdoMet synthetase. Preincubation of enzyme fraction with L-Methionine failed to activate or stabilize the activity of AdoMet synthetase. Three isozymes of AdoMet synthetase were physically separated by DE-52 ion-exchange chromatography. One of the isozymes of AdoMet synthetase has been purified (1529-fold) to electrophoretic homogeneity by resorting to phenyl Sepharose and ATP Sepharose affinity chromatography. The purified enzyme catalyzed the synthesis of S-adenosylmethionine and also exhibited tripolyphosphatase activity. The reaction product of the purified enzyme was chemically and enzymatically characterized as S-adenosylmethionine. The molecular weight of the native enzyme is 174,000 and that of its subunit is 84,000 as determined on SDS-PAGE. Thus the native enzyme seems to be dimeric in nature. PMID:1648405

  8. 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.

  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. 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.

  11. 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…

  12. 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.

  13. Distortion correction for the orthogonally-splitting-imaging pose sensor

    NASA Astrophysics Data System (ADS)

    Yang, Qian; Sun, Chang-ku; Wang, Peng; Li, Wen-qiang; Liu, Xin-tong

    2015-06-01

    The orthogonally-splitting-imaging pose sensor utilizes not only large field of view spherical lenses but also two sets of cylindrical ones to realize the high-speed, high-precision and wide-field pose measurement. Notable distortion, however, results from the wide-field lenses at the same time. Therefore, to obtain the best performance of the camera model, a distortion correction method is proposed in this paper, which combines the advantages of the high-stability of the Least Square fittings based on the orthogonal polynomials and the independence of the distortion correction based on the cross-ratio invariability. In this way, the ill-conditioned fitting matrix as well as the iteration and optimization procedures in solving extrinsic and intrinsic parameters can be avoided. Due to the wide fitness of the cross-ratio invariability and the orthogonal polynomials, this distortion correction technique is also suitable to other optical set up with different imaging structure. The experiment results that the corrected grids have superior precision and reliability with their original slopes demonstrate that the distortion model on the basis of orthogonal polynomial is validated and that the distortion correction is effective.

  14. Complex Exceptional Orthogonal Polynomials and Quasi-invariance

    NASA Astrophysics Data System (ADS)

    Haese-Hill, William A.; Hallnäs, Martin A.; Veselov, Alexander P.

    2016-05-01

    Consider the Wronskians of the classical Hermite polynomials H_{λ, l}(x):=Wr(H_l(x),H_{k_1}(x),ldots,H_{k_n}(x)), quad l in Z_{≥0}{setminus} {k_1,ldots,k_n}, where {k_i=λ_i+n-i, i=1,ldots, n} and {λ=(λ_1, ldots, λ_n)} is a partition. Gómez-Ullate et al. showed that for a special class of partitions the corresponding polynomials are orthogonal and dense among all polynomials with respect to a certain inner product, but in contrast to the usual case have some degrees missing (so-called exceptional orthogonal polynomials). We generalise their results to all partitions by considering complex contours of integration and non-positive Hermitian products. The corresponding polynomials are orthogonal and dense in a finite-codimensional subspace of {C[x]} satisfying certain quasi-invariance conditions. A Laurent version of exceptional orthogonal polynomials, related to monodromy-free trigonometric Schrödinger operators, is also presented.

  15. Alternative Kondo breakdown mechanism: Orbital-selective orthogonal metal transition

    NASA Astrophysics Data System (ADS)

    Zhong, Yin; Liu, Ke; Wang, Yong-Qiang; Luo, Hong-Gang

    2012-09-01

    In a recent paper of Nandkishore, Metlitski, and Senthil [Phys. Rev. B1098-012110.1103/PhysRevB.86.045128 86, 045128 (2012)], a concept of orthogonal metal has been introduced to reinterpret the disordered state of slave-spin representation in the Hubbard model as an exotic gapped metallic state. We extend this concept to study the corresponding quantum phase transition in the extended Anderson lattice model. It is found that the disordered state of slave spins in this model is an orbital-selective orthogonal metal, a generalization of the concept of the orthogonal metal in the Hubbard model. The quantum critical behaviors are multiscale and dominated by a z=3 and z=2 critical modes in the high- and low-temperature regime, respectively. Such behaviors are obviously in contrast to the naive expectation in the Hubbard model. The result provides alternative Kondo breakdown mechanism for heavy fermion compounds underlying the physics of the orbital-selective orthogonal metal in the disordered state, which is different from the conventional Kondo breakdown mechanism with the fractionalized Fermi-liquid picture. This work is expected to be useful in understanding the quantum criticality happening in some heavy fermion materials and other related strongly correlated systems.

  16. A Monotonically Convergent Algorithm for Orthogonal Congruence Rotation.

    ERIC Educational Resources Information Center

    Kiers, Henk A. L.; Groenen, Patrick

    1996-01-01

    An iterative majorization algorithm is proposed for orthogonal congruence rotation that is guaranteed to converge from every starting point. In addition, the algorithm is easier to program than the algorithm proposed by F. B. Brokken, which is not guaranteed to converge. The derivation of the algorithm is traced in detail. (SLD)

  17. 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

  18. Divergent anticodon recognition in contrasting glutamyl-tRNA synthetases.

    PubMed

    Lee, Joohee; Hendrickson, Tamara L

    2004-12-10

    The pathogenic bacterium Helicobacter pylori utilizes two essential glutamyl-tRNA synthetases (GluRS1 and GluRS2). These two enzymes are closely related in evolution and yet they aminoacylate contrasting tRNAs. GluRS1 is a canonical discriminating GluRS (D-GluRS) that biosynthesizes Glu-tRNA(Glu) and cannot make Glu-tRNA(Gln). In contrast, GluRS2 is non-canonical as it is only essential for the production of misacylated Glu-tRNA(Gln). The co-existence and evident divergence of these two enzymes was capitalized upon to directly examine how GluRS2 acquired tRNA(Gln) specificity. One key feature that distinguishes tRNA(Glu) from tRNA(Gln) is the third position in the anticodon of each tRNA (C36 versus G36, respectively). By comparing sequence alignments of different GluRSs, including GluRS1s and GluRS2s, to the crystal structure of the Thermus thermophilus D-GluRS:tRNA(Glu) complex, a divergent pattern of conservation in enzymes that aminoacylate tRNA(Glu)versus those specific for tRNA(Gln) emerged and was experimentally validated. In particular, when an arginine conserved in discriminating GluRSs and GluRS1s was inserted into Hp GluRS2 (Glu334Arg GluRS2), the catalytic efficiency of the mutant enzyme (k(cat)/K(Mapp)) was reduced by approximately one order of magnitude towards tRNA(Gln). However, this mutation did not introduce activity towards tRNA(Glu). In contrast, disruption of a glycine that is conserved in all GluRS2s but not in other GluRSs (Gly417Thr GluRS2) generated a mutant GluRS2 with weak activity towards tRNA(Glu1). Synergy between these two mutations was observed in the double mutant (Glu334Arg/Gly417Thr GluRS2), which specifically and more robustly aminoacylates tRNA(Glu1) instead of tRNA(Gln). As GluRS1 and GluRS2 are related by an apparent gene duplication event, these results demonstrate that we can experimentally map critical evolutionary events in the emergence of new tRNA specificities. PMID:15561136

  19. Crystal structure of NH3-dependent NAD+ synthetase from Bacillus subtilis.

    PubMed Central

    Rizzi, M; Nessi, C; Mattevi, A; Coda, A; Bolognesi, M; Galizzi, A

    1996-01-01

    NAD+ synthetase catalyzes the last step in the biosynthesis of nicotinamide adenine dinucleotide. The three-dimensional structure of NH3-dependent NAD+ synthetase from Bacillus subtilis, in its free form and in complex with ATP, has been solved by X-ray crystallography (at 2.6 and 2.0 angstroms resolution, respectively) using a combination of multiple isomorphous replacement and density modification techniques. The enzyme consists of a tight homodimer with alpha/beta subunit topology. The catalytic site is located at the parallel beta-sheet topological switch point, where one AMP molecule, one pyrophosphate and one Mg2+ ion are observed. Residue Ser46, part of the neighboring 'P-loop', is hydrogen bonded to the pyrophosphate group, and may play a role in promoting the adenylation of deamido-NAD+ during the first step of the catalyzed reaction. The deamido-NAD+ binding site, located at the subunit interface, is occupied by one ATP molecule, pointing towards the catalytic center. A conserved structural fingerprint of the catalytic site, comprising Ser46, is very reminiscent of a related protein region observed in glutamine-dependent GMP synthetase, supporting the hypothesis that NAD+ synthetase belongs to the newly discovered family of 'N-type' ATP pyrophosphatases. Images PMID:8895556

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

    PubMed

    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

    2015-12-01

    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

  1. The parallel and convergent universes of polyketide synthases and nonribosomal peptide synthetases.

    PubMed

    Cane, D E; Walsh, C T

    1999-12-01

    Polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) catalyze chain elongation from simple building blocks to create a diverse array of natural products. PKS and NRPS proteins share striking architectural and organizational similarities that can be exploited to generate entirely new natural products. PMID:10631508

  2. 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

  3. Structure of Pyrrolysyl-tRNA Synthetase, an Archaeal Enzyme for Genetic Code Innovation

    SciTech Connect

    Kavran,J.; Gundllapalli, S.; O'Donoghue, P.; Englert, M.; Soll, D.; Steitz, T.

    2007-01-01

    Pyrrolysine (Pyl), the 22nd natural amino acid and genetically encoded by UAG, becomes attached to its cognate tRNA by pyrrolysyl-tRNA synthetase (PylRS). We have determined three crystal structures of the Methanosarcina mazei PylRS complexed with either AMP-PNP, Pyl-AMP plus pyrophosphate, or the Pyl analogue N-e-[(cylopentyloxy)carbonyl]-l-lysine plus ATP. The structures reveal that PylRS utilizes a deep hydrophobic pocket for recognition of the Pyl side chain. A comparison of these structures with previously determined class II tRNA synthetase complexes illustrates that different substrate specificities derive from changes in a small number of residues that form the substrate side-chain-binding pocket. The knowledge of these structures allowed the placement of PylRS in the aminoacyl-tRNA synthetase (aaRS) tree as the last known synthetase that evolved for genetic code expansion, as well as the finding that Pyl arose before the last universal common ancestral state. The PylRS structure provides an excellent framework for designing new aaRSs with altered amino acid specificity.

  4. Phosphoribosyl pyrophosphate synthetase activity affects growth and riboflavin production in Ashbya gossypii

    PubMed Central

    Jiménez, Alberto; Santos, María A; Revuelta, José L

    2008-01-01

    Background Phosphoribosyl pyrophosphate (PRPP) is a central compound for cellular metabolism and may be considered as a link between carbon and nitrogen metabolism. PRPP is directly involved in the de novo and salvage biosynthesis of GTP, which is the immediate precursor of riboflavin. The industrial production of this vitamin using the fungus Ashbya gossypii is an important biotechnological process that is strongly influenced by substrate availability. Results Here we describe the characterization and manipulation of two genes of A. gossypii encoding PRPP synthetase (AGR371C and AGL080C). We show that the AGR371C and AGL080C gene products participate in PRPP synthesis and exhibit inhibition by ADP. We also observed a major contribution of AGL080C to total PRPP synthetase activity, which was confirmed by an evident growth defect of the Δagl080c strain. Moreover, we report the overexpression of wild-type and mutant deregulated isoforms of Agr371cp and Agl080cp that significantly enhanced the production of riboflavin in the engineered A. gossypii strains. Conclusion It is shown that alterations in PRPP synthetase activity have pleiotropic effects on the fungal growth pattern and that an increase in PRPP synthetase enzymatic activity can be used to enhance riboflavin production in A. gossypii. PMID:18782443

  5. Characterization of recombinant glutamine synthetase from the hyperthermophilic archaeon Pyrococcus sp. strain KOD1.

    PubMed Central

    Adul Rahman, R N; Jongsareejit, B; Fujiwara, S; Imanaka, T

    1997-01-01

    The glnA gene encoding glutamine synthetase was cloned from the hyperthermophilic archaeon Pyrococcus sp. strain KOD1, and its nucleotide sequence was determined. The glnA gene was expressed in Escherichia coli ME8459 (glnA mutant strain), and the protein was purified to homogeneity and shown to be functional in a dodecameric from (637,000 Da), exhibiting both transferase and synthetase activities. However, kinetic studies indicated that the enzyme possessed low biosynthetic activity, suggesting that the reaction was biased towards glutamate production. The optimum temperature for both activities was 60 degrees C, which was lower than the optimal growth temperature of KOD1. Recombinant KOD1 GlnA exhibited different optimum pHs depending on the reaction employed (pH 7.8 for the synthetase reaction and pH 7.2 for the transferase reaction). Of the various nucleoside triphosphates tested, GTP as well as ATP was involved in the synthetase reaction. PMID:9172372

  6. 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

  7. Augmenting ureagenesis in patients with partial carbamyl phosphate synthetase 1deficiency with N-carbamylglutamate

    PubMed Central

    Ah Mew, Nicholas; McCarter, Robert; Daikhin, Yevgeny; Lichter, Uta; Nissim, Ilana; Yudkoff, Marc; Tuchman, and Mendel

    2014-01-01

    Identical studies employing stable isotopes were performed before and after a 3-day trial of oral N-carbamylglutamate (NCG) in 5 subjects with late onset carbamyl phosphate synthetase deficiency. NCG augmented ureagenesis and decreased plasma ammonia in 4 of 5 subjects. There was marked improvement in nitrogen metabolism with long-term NCG administration in one subject. PMID:24880889

  8. Structures of two distinct conformations of holo-non-ribosomal peptide synthetases.

    PubMed

    Drake, Eric J; Miller, Bradley R; Shi, Ce; Tarrasch, Jeffrey T; Sundlov, Jesse A; Allen, C Leigh; Skiniotis, Georgios; Aldrich, Courtney C; Gulick, Andrew M

    2016-01-14

    Many important natural products are produced by multidomain non-ribosomal peptide synthetases (NRPSs). During synthesis, intermediates are covalently bound to integrated carrier domains and transported to neighbouring catalytic domains in an assembly line fashion. Understanding the structural basis for catalysis with non-ribosomal peptide synthetases will facilitate bioengineering to create novel products. Here we describe the structures of two different holo-non-ribosomal peptide synthetase modules, each revealing a distinct step in the catalytic cycle. One structure depicts the carrier domain cofactor bound to the peptide bond-forming condensation domain, whereas a second structure captures the installation of the amino acid onto the cofactor within the adenylation domain. These structures demonstrate that a conformational change within the adenylation domain guides transfer of intermediates between domains. Furthermore, one structure shows that the condensation and adenylation domains simultaneously adopt their catalytic conformations, increasing the overall efficiency in a revised structural cycle. These structures and the single-particle electron microscopy analysis demonstrate a highly dynamic domain architecture and provide the foundation for understanding the structural mechanisms that could enable engineering of novel non-ribosomal peptide synthetases. PMID:26762461

  9. Gain-of-Function Mutational Activation of Human tRNA Synthetase Procytokine

    PubMed Central

    Yang, Xiang-Lei; Kapoor, Mili; Otero, Francella J.; Slike, Bonnie M.; Tsuruta, Hiro; Frausto, Ricardo; Bates, Alison; Ewalt, Karla L.; Cheresh, David A.; Schimmel, Paul

    2008-01-01

    Summary Disease-causing mutations occur in genes for aminoacyl tRNA synthetases. That some mutations are dominant suggests a gain-of-function. Native tRNA synthetases, like TyrRS and TrpRS, 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 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. PMID:18096501

  10. DISTRIBUTION OF THE GLUTAMINE SYNTHETASE ISOZYME GSP1 IN MAIZE (ZEA MAYS)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Higher plants contain families of glutamine synthetase (GS) isozymes. In maize (Zea mays L.), GSp1, the predominant GS isozyme of the developing kernel, is abundant in the pedicel and pericarp, but absent from the endosperm and embryo. Determination of GSp1 tissue distribution in vegetative tissue...

  11. Synthesis and activities of branched-chain aminoacyl-tRNA synthetases in threonine deaminase mutants of Escherichia coli.

    PubMed Central

    Williams, A L; Whitfield, S M; Williams, L S

    1978-01-01

    Valyl-, isoleucyl-, and leucyl-tRNA synthetase activities were examined in an Escherichia coli K-12 strain that possessed a deletion of three genes of the ilv gene cluster, ilvD, A, and C, and in a strain with the same deletion that also carried the lambdadilvCB bacteriophage. It was observed that the branched-chain tRNA synthetase activities of both strains were considerably less than those of the normal strain during growth in unrestricted medium. Furthermore, during an isoleucine limitation, there was a further reduction in isoleucyl-tRNA synthetase activity and an absence of the isoleucine-mediated derepression of valyl-tRNA synthetase formation in both of these mutants, as compared with the normal strain. In addition, it was observed that these branched-chain synthetase activities were reduced in steady-state cultures of several ilvA point mutants. However, upon the introduction of the ilv operon to these ilvA mutants by use of lambda bacteriophage, there was a specific increase in the branched-chain synthetase activities to levels comparable to those of the normal strain. These results support our previous findings that the stability and repression control of synthesis of these synthetases require some product(s) missing in the ilvDAC deletion strain and strongly suggest this component is some form of the ilvA gene product, threonine deaminase. PMID:348689

  12. Inactivation and dissociation of S-adenosylmethionine synthetase by modification of sulfhydryl groups and its possible occurrence in cirrhosis.

    PubMed

    Corrales, F; Cabrero, C; Pajares, M A; Ortiz, P; Martin-Duce, A; Mato, J M

    1990-02-01

    Catalytically active human and rat liver S-adenosylmethionine synthetase exists mainly in tetramer and dimer form. In liver biopsy samples from cirrhotic patients a marked reduction in total S-adenosylmethionine synthetase activity and a specific loss of the tetrameric form of the enzyme exist. We have investigated the possible role of sulfhydryl groups in maintaining the structure and activity of S-adenosylmethionine synthetase. Both forms of S-adenosylmethionine synthetase are rapidly inactivated by N-ethylmaleimide, and the loss of enzyme activity correlates with the incorporation of approximately 2 moles N-ethylmaleimide per mole of subunit. In addition, reaction with N-ethylmaleimide resulted in displacement of the tetramer-dimer equilibrium of the enzyme toward the dimer, but no monomer was detected under these conditions. A catalytically active monomeric S-adenosylmethionine synthetase was detected in the cytosolic extract from a liver biopsy sample from a cirrhotic patient, supporting our model for the structure of S-adenosylmethionine synthetase. Because treatment of S-adenosylmethionine synthetase with N-ethylmaleimide resembles the situation of this enzyme in cirrhotic patients, it is proposed that impaired protection of the enzyme from oxidizing agents caused by a decreased synthesis of glutathione can explain the diminished synthesis of S-adenosylmethionine in liver cirrhosis. PMID:2307400

  13. An example of non-conservation of oligomeric structure in prokaryotic aminoacyl-tRNA synthetases. Biochemical and structural properties of glycyl-tRNA synthetase from Thermus thermophilus.

    PubMed

    Mazauric, M H; Reinbolt, J; Lorber, B; Ebel, C; Keith, G; Giegé, R; Kern, D

    1996-11-01

    Glycyl-tRNA synthetase (Gly-tRNA synthetase) from Thermus thermophilus was purified to homogeneity and with high yield using a five-step purification procedure in amounts sufficient to solve its crystallographic structure [Logan, D.T., Mazauric, M.-H., Kern, D. & Moras, D. (1995) EMBO J. 14, 4156-4167]. Molecular-mass determinations of the native and denatured protein indicate an oligomeric structure of the alpha 2 type consistent with that found for eukaryotic Gly-tRNA synthetases (yeast and Bombyx mori), but different from that of Gly-tRNA synthetases from mesophilic prokaryotes (Escherichia coli and Bacillus brevis) which are alpha 2 beta 2 tetramers. N-terminal sequencing of the polypeptide chain reveals significant identity, reaching 50% with those of the eukaryotic enzymes (B. mori, Homo sapiens, yeast and Caenorhabditis elegans) but no significant identity was found with both alpha and beta chains of the prokaryotic enzymes (E. coli, Haemophilus influenzae and Coxiella burnetii) albeit the enzyme is deprived of the N-terminal extension characterizing eukaryotic synthetases. Thus, the thermophilic Gly-tRNA synthetase combines strong structural homologies of eukaryotic Gly-tRNA synthetases with a feature of prokaryotic synthetases. Heat-stability measurements show that this synthetase keeps its ATP-PPi exchange and aminoacylation activities up to 70 degrees C. Glycyladenylate strongly protects the enzyme against thermal inactivation at higher temperatures. Unexpectedly, tRNA(Gly) does not induce protection. Cross-aminoacylations reveal that the thermophilic Gly-tRNA synthetase charges heterologous E. coli tRNA(gly(GCC)) and tRNA(Gly(GCC)) and yeast tRNA(Gly(GCC)) as efficiently as T. thermophilus tRNA(Gly). All these aminoacylation reactions are characterized by similar activation energies as deduced from Arrhenius plots. Therefore, contrary to the E. coli and H. sapiens Gly-tRNA synthetases, the prokaryotic thermophilic enzyme does not possess a strict

  14. Helicobacter pylori Glutamine Synthetase Lacks Features Associated with Transcriptional and Posttranslational Regulation

    PubMed Central

    Garner, Rachel M.; Fulkerson, John; Mobley, Harry L. T.

    1998-01-01

    Helicobacter pylori urease, produced in abundance, is indispensable for the survival of H. pylori in animal hosts. Urea is hydrolyzed by the enzyme, resulting in the liberation of excess ammonia, some of which neutralizes gastric acid. The remaining ammonia is assimilated into protein by glutamine synthetase (EC 6.3.1.2), which catalyzes the reaction: NH3 + glutamate + ATP→glutamine + ADP + Pi. We hypothesized that glutamine synthetase plays an unusually critical role in nitrogen assimilation by H. pylori. We developed a phenotypic screen to isolate genes that contribute to the synthesis of a catalytically active urease. Escherichia coli SE5000 transformed with plasmid pHP808 containing the entire H. pylori urease gene cluster was cotransformed with a pBluescript plasmid library of the H. pylori ATCC 43504 genome. A weakly urease-positive 9.4-kb clone, pUEF728, was subjected to nucleotide sequencing. Among other genes, the gene for glutamine synthetase was identified. The complete 1,443-bp glnA gene predicts a polypeptide of 481 amino acid residues with a molecular weight of 54,317; this was supported by maxicell analysis of cloned glnA expressed in E. coli. The top 10 homologs were all bacterial glutamine synthetases, including Salmonella typhimurium glnA. The ATP-binding motif GDNGSG (residues 272 to 277) of H. pylori GlnA exactly matched and aligned with the sequence in 8 of the 10 homologs. The adenylation site found in the top 10 homologs (consensus sequence, NLYDLP) is replaced in H. pylori by NLFKLT (residues 405 to 410). Since the Tyr (Y) residue is the target of adenylation and since the H. pylori glutamine synthetase lacks that residue in four strains examined, we conclude that no adenylation occurs within this motif. Cloned H. pylori glnA complemented a glnA mutation in E. coli, and GlnA enzyme activity could be measured spectrophotometrically. In an attempt to produce a GlnA-deficient mutant of H. pylori, a kanamycin resistance cassette was cloned

  15. 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

  16. 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.

  17. Primary structure of histidine-tRNA synthetase and characterization of hisS transcripts.

    PubMed

    Freedman, R; Gibson, B; Donovan, D; Biemann, K; Eisenbeis, S; Parker, J; Schimmel, P

    1985-08-25

    Histidine-tRNA synthetase is one of the smallest bacterial aminoacyl-tRNA synthetases. It is less than one-half the size of the largest aminoacyl-tRNA synthetases. The entire nucleotide sequence of the Escherichia coli hisS locus was determined. The coding region is comprised of 424 codons, and the sequence was determined for 200 nucleotides on the 5'- and 3'-sides of the coding region. The translated nucleotide sequence was confirmed extensively by independent amino acid sequence information obtained by Edman degradations of purified peptides and by measurements of peptide masses by fast atom bombardment mass spectrometry. A significant sequence alignment of four bacterial aminoacyl-tRNA synthetases was reported recently (Webster, T., Tsai, H., Kula, M., Mackie, G., and Schimmel, P. (1984) Science 226, 1315-1317). Although the four enzymes vary considerably in length, this match occurs within the first 100 amino acids of each of the four enzymes and is in the segment believed to be part of the catalytic core. But no strong alignment could be found of the histidine sequence with these four tRNA synthetase sequences. This enzyme may be derived, therefore, from a different progenitor. Previous work suggested that three places in the hisS 5'-noncoding sequence could be promoter sites for RNA polymerase (Eisenbeis, S. J., and Parker, J. (1982) Gene 18, 107-114). We detected a 1400-nucleotide RNA species by RNA blot analysis with a hisS-specific probe. S1 nuclease mapping demonstrated a 5'-end to the RNA species occurs at -67 +/- 1, relative to the first nucleotide of the coding region. This position coincides with the predicted start site for transcription from one of the previously proposed promoter sites. PMID:2991272

  18. Properties and substrate specificities of the phenylalanyl-transfer-ribonucleic acid synthetases of Aesculus species

    PubMed Central

    Anderson, J. W.; Fowden, L.

    1970-01-01

    1. Phenylalanyl-tRNA synthetases have been partially purified from cotyledons of seeds of Aesculus californica, which contains 2-amino-4-methylhex-4-enoic acid, and from four other species of Aesculus that do not contain this amino acid. The A. californica preparation was free from other aminoacyl-tRNA synthetases, and the contaminating synthetase activity in preparations from A. hippocastanum was decreased to acceptable limits by conducting assays of pyrophosphate exchange activity in 0.5m-potassium chloride. 2. The phenylalanyl-tRNA synthetase from each species activated 2-amino-4-methylhex-4-enoic acid with Km 30–40 times that for phenylalanine. The maximum velocity for 2-amino-4-methylhex-4-enoic acid was only 30% of that for phenylalanine with the A. californica enzyme, but the maximum velocities for the two substrates were identical for the other four species. 3. 2-Amino-4-methylhex-4-enoic acid was not found in the protein of A. californica, so discrimination against this amino acid probably occurs in the step of transfer to tRNA, though subcellular localization, or subsequent steps of protein synthesis could be involved. 4. Crotylglycine, methallylglycine, ethallylglycine, 2-aminohex-4,5-dienoic acid, 2-amino-5-methylhex-4-enoic acid, 2-amino-4-methylhex-4-enoic acid, β-(thien-2-yl)alanine, β-(pyrazol-1-yl)alanine, phenylserine and m-fluorophenylalanine were substrates for pyrophosphate exchange catalysed by the phenylalanyl-tRNA synthetases of A. californica or A. hippocastanum. Allylglycine, phenylglycine and 2-amino-4-phenylbutyric acid were inactive. PMID:5493504

  19. 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.

  20. 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

  1. 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

  2. Orthogonal canonical forms for second-order systems

    NASA Technical Reports Server (NTRS)

    Williams, Trevor; Laub, Alan

    1989-01-01

    The authors prove that a linear second-order system with arbitrary damping cannot be reduced to Hessenberg-triangular form by means of orthogonal transformations, while this reduction is always possible for the modal damping commonly assumed for models of flexible structures. The type of canonical form obtainable by means of orthogonal transformations acting on a second-order system is heavily dependent on the type of damping considered. If the damping matrix is merely positive semi-definite symmetric, it is generally not possible to obtain a reduction to Hessenberg-triangular form, while this reduction is trivial for zero or Rayleigh damping. If damping is modal, however, as is commonly assumed in structural models, the reduction exists and is nontrivial. Furthermore, reduction to triangular second-order Schur form is always possible for such damping: this canonical form appears likely to have applications to second-order system theory.

  3. 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. PMID:25791164

  4. Orthogonal bone cutting: saw design and operating characteristics.

    PubMed

    Krause, W R

    1987-08-01

    The cutting process of orthopaedic bone saws was considered as orthogonal (two-dimensional) cutting for determination of the horizontal and vertical force components of single edge cutting tools with rake angles of 0 to -30 degrees. The Merchant analysis for orthogonal cutting was used to determine the resultant force and other force and work relationships. The effect of an imposed lateral vibration on the cutting tool was also investigated. The results of the tests indicated a strong interaction between the measured and derived forces with the rake angle and feed velocity. It was concluded that to reduce the cutting forces and work expenditure, a negative rake angle between 0 and -10 degrees, high feed velocity, and an imposed lateral vibration provided the greatest reduction in force and energy expenditure. PMID:3657116

  5. External orthogonality in subsystem time-dependent density functional theory.

    PubMed

    Chulhai, Dhabih V; Jensen, Lasse

    2016-08-01

    Subsystem density functional theory (subsystem DFT) is a DFT partitioning method that is exact in principle, but depends on approximations to the kinetic energy density functional (KEDF). One may avoid the use of approximate KEDFs by ensuring that the inter-subsystem molecular orbitals are orthogonal, termed external orthogonality (EO). We present a method that extends a subsystem DFT method, that includes EO, into the time-dependent DFT (TDDFT) regime. This method therefore removes the need for approximations to the kinetic energy potential and kernel, and we show that it can accurately reproduce the supermolecular TDDFT results for weakly and strongly coupled subsystems, and for systems with strongly overlapping densities (where KEDF approximations traditionally fail). PMID:26932176

  6. Orthogonal polarization Mirau interferometer using reflective-type waveplate.

    PubMed

    Tapilouw, Abraham Mario; Chen, Liang-Chia; Jen, Yi-Jun; Lin, Shyh-Tsong; Yeh, Sheng-Lih

    2013-07-15

    This work proposes an orthogonal polarization Mirau interferometry using a reflective-type waveplate to generate different polarization orientations for broadband white light interferometry. The reflective-type half-waveplate is employed as the reference arm of the Mirau interferometer to convert polarization and it generates a reference light with an orientation orthogonal to the object light. An advantage of the proposed interferometer is its ability to control the ratio of light intensity between the object and reference arms to maximize the interferometric fringe contrast. Better, more accurate calibration of standard step height has been achieved by the developed interferometer, which also can measure solder bumps that traditional Mirau interferometers usually cannot measure. PMID:23939094

  7. 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

  8. 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.

  9. Orthogonal polynomial interpretation of Δ-Toda equations

    NASA Astrophysics Data System (ADS)

    Area, I.; Branquinho, A.; Foulquié Moreno, A.; Godoy, E.

    2015-10-01

    In this paper a discretization of Toda equations is analyzed. The correspondence between these Δ-Toda equations for the coefficients of the Jacobi operator and its resolvent function is established. It is shown that the spectral measure of these operators evolve in t like {(1+x)}1-t {{d}}μ (x) where {{d}}μ is a given positive Borel measure. The Lax pair for the Δ-Toda equations is derived and characterized in terms of linear functionals, where orthogonal polynomials which satisfy an Appell condition with respect to the forward difference operator Δ appear in a natural way. In order to illustrate the results of the paper we work out two examples of Δ-Toda equations related with Jacobi and Laguerre orthogonal polynomials.

  10. 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.

  11. 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.

  12. Star polymer synthesis viaλ-orthogonal photochemistry.

    PubMed

    Hiltebrandt, Kai; Kaupp, Michael; Molle, Edgar; Menzel, Jan P; Blinco, James P; Barner-Kowollik, Christopher

    2016-08-01

    We introduce a light induced sequence enabling λ-orthogonal star polymer formation via an arms-first approach, based on an α,ω-functional polymer carrying tetrazole and o-methyl benzaldehyde moieties, which upon irradiation can readily undergo cycloaddition with a trifunctional maleimide core. Depending on the wavelength, the telechelic strand can be attached to the core at either photo-reactive end. PMID:27378494

  13. Parsimonious extreme learning machine using recursive orthogonal least squares.

    PubMed

    Wang, Ning; Er, Meng Joo; Han, Min

    2014-10-01

    Novel constructive and destructive parsimonious extreme learning machines (CP- and DP-ELM) are proposed in this paper. By virtue of the proposed ELMs, parsimonious structure and excellent generalization of multiinput-multioutput single hidden-layer feedforward networks (SLFNs) are obtained. The proposed ELMs are developed by innovative decomposition of the recursive orthogonal least squares procedure into sequential partial orthogonalization (SPO). The salient features of the proposed approaches are as follows: 1) Initial hidden nodes are randomly generated by the ELM methodology and recursively orthogonalized into an upper triangular matrix with dramatic reduction in matrix size; 2) the constructive SPO in the CP-ELM focuses on the partial matrix with the subcolumn of the selected regressor including nonzeros as the first column while the destructive SPO in the DP-ELM operates on the partial matrix including elements determined by the removed regressor; 3) termination criteria for CP- and DP-ELM are simplified by the additional residual error reduction method; and 4) the output weights of the SLFN need not be solved in the model selection procedure and is derived from the final upper triangular equation by backward substitution. Both single- and multi-output real-world regression data sets are used to verify the effectiveness and superiority of the CP- and DP-ELM in terms of parsimonious architecture and generalization accuracy. Innovative applications to nonlinear time-series modeling demonstrate superior identification results. PMID:25291736

  14. Time-resolved proper orthogonal decomposition of liquid jet dynamics

    NASA Astrophysics Data System (ADS)

    Arienti, Marco; Soteriou, Marios C.

    2009-11-01

    New insight into the mechanism of liquid jet in crossflow atomization is provided by an analysis technique based on proper orthogonal decomposition and spectral analysis. Data are provided in the form of high-speed videos of the jet near field from experiments over a broad range of injection conditions. For each condition, proper orthogonal modes (POMs) are generated and ordered by intensity variation relative to the time average. The feasibility of jet dynamics reduction by truncation of the POM series to the first few modes is then examined as a function of crossflow velocity for laminar and turbulent liquid injection. At conditions where the jet breaks up into large chunks of liquid, the superposition of specific orthogonal modes is observed to track long waves traveling along the liquid column. The temporal coefficients of these modes can be described as a bandpass spectrum that shifts toward higher frequencies as the crossflow velocity is increased. The dynamic correlation of these modes is quantified by their cross-power spectrum density. Based on the frequency and wavelength extracted from the videos, the observed traveling waves are linked to the linearly fastest growing wave of Kelvin-Helmholtz instability. The gas boundary layer thickness at the gas-liquid shear layer emerges at the end of this study as the dominant length scale of jet dynamics at moderate Weber numbers.

  15. 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

  16. 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

  17. 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. PMID:18467751

  18. Orthogonality breaking through few-mode optical fiber.

    PubMed

    Parnet, Francois; Fade, Julien; Alouini, Mehdi

    2016-04-01

    Polarization sensing and imaging through optical fibers is a technological challenge motivated by promising applications for in vivo, in situ polarimetric endoscopy for biomedical diagnosis. Among the recent approaches proposed to solve this issue, the depolarization/dichroism sensing by polarization orthogonality breaking (DSOB) technique was shown to perform remotely through single-mode optical fibers for depolarization/diattenuation measurements. In this article, we investigate the applicability of such a technique in slightly multimode waveguides. Through theoretical modeling and numerical simulations, we evidence the conditions required for the polarization orthogonality to be preserved after propagation in a few-mode fiber, notably in terms of detection geometry of the spatial modes. Original experiments realized in few-mode fibers both in transmission and reflection configurations are also reported and validate the theoretical predictions. These results allow us to analyze the influence of the experimental parameters, such as detection geometry, sample tilt, or fiber length, on orthogonality preservation and on the measurement dynamics of the DSOB technique in slightly multimode waveguides. PMID:27139651

  19. Multilayer block copolymer meshes by orthogonal self-assembly

    NASA Astrophysics Data System (ADS)

    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.

  20. Orthogonal reference pattern multiplexing for collinear holographic data storage.

    PubMed

    Cao, Liangcai; Liu, Jinqiu; Li, Jianhua; He, Qingsheng; Jin, Guofan

    2014-01-01

    An orthogonal reference pattern multiplexing (ORPM) method for collinear holographic data storage (CHDS) is investigated to increase the data storage density and realize parallel optical image superimposition. Holograms are multiplexed in the same volume of the recording medium with multiple orthogonal reference patterns (RPs). The physical principle of this method is analyzed based on scalar diffraction theory. The orthogonal condition of the RPs is derived in order to suppress the interpage cross talk. The parameters of the radial-line RP have significant influence on the signal-to-noise ratio (SNR) of the reconstructed data page. They are optimized to reduce the intrapage cross talk. With a random binary phase mask (RBPM) located closely before the spatial light modulator, SNR of the reconstructed data page is seven times the SNR without the RBPM. Three data pages are multiplexed in the same volume of the medium using the ORPM method. The reconstructed data pages for the CHDS system show the effectiveness of the RBPM in suppressing the intrapage and interpage cross talk. PMID:24513981

  1. 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 .

  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. Minimal parameter solution of the orthogonal matrix differential equation

    NASA Technical Reports Server (NTRS)

    Baritzhack, 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.

  4. 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.

  5. Coherent vorticity extraction in resistive drift-wave turbulence: Comparison of orthogonal wavelets versus proper orthogonal decomposition

    NASA Astrophysics Data System (ADS)

    Futatani, Shimpei; Bos, Wouter J. T.; del-Castillo-Negrete, Diego; Schneider, Kai; Benkadda, Sadruddin; Farge, Marie

    2011-03-01

    We assess two techniques for extracting coherent vortices out of turbulent flows: the wavelet based Coherent Vorticity Extraction (CVE) and the Proper Orthogonal Decomposition (POD). The former decomposes the flow field into an orthogonal wavelet representation and subsequent thresholding of the coefficients allows one to split the flow into organized coherent vortices with non-Gaussian statistics and an incoherent random part which is structureless. POD is based on the singular value decomposition and decomposes the flow into basis functions which are optimal with respect to the retained energy for the ensemble average. Both techniques are applied to direct numerical simulation data of two-dimensional drift-wave turbulence governed by Hasegawa-Wakatani equation, considering two limit cases: the quasi-hydrodynamic and the quasi-adiabatic regimes. The results are compared in terms of compression rate, retained energy, retained enstrophy and retained radial flux, together with the enstrophy spectrum and higher order statistics.

  6. Active site nanospace of aminoacyl tRNA synthetase: difference between the class I and class II synthetases.

    PubMed

    Dutta, Saheb; Choudhury, Kaberi; Banik, Sindrila Dutta; Nandi, Nilashis

    2014-03-01

    The present work is aimed at understanding the origin of the difference in the molecular organization of the active site nanospaces of the class I and class II aminoacyl tRNA synthetases (aaRSs) which are tunnel-like structures. The active site encloses the cognate amino acid (AA) and the adenosine triphosphate (ATP) to carry out aminoacylation reaction. Comparison of the structures of the active site of the class I and class II (aaRSs) shows that the nanodimensional tunnels are curved in opposite directions in the two classes. We investigated the origin of this difference using quantum mechanical computation of electrostatic potential (ESP) of substrates, surrounding residues and ions, using Atoms in Molecule (AIM) Theory and charge population analysis. We show that the difference is principally due to the variation in the spatial charge distribution of ATP in the two classes which correspond to extended and bent conformations of ATP. The present computation shows that the most feasible pathway for nucleophilic attack to alphaP is oppositely directed for class I and class II aaRSs. The available crystal structures show that the cognate AA is indeed located along the channel favorable for nucleophilic attack as predicted by the ESP analysis. It is also shown that the direction of the channel changes its orientation when the orientation of ATP is changed from extended to a bent like structure. We further used the AIM theory to confirm the direction of the approach of AA in each case and the results corroborate the results from the ESP analysis. The opposite curvatures of the active site nanospaces in class I and class II aaRSs are related with the influence of the charge distributions of the extended and bent conformations of ATP, respectively. The results of the computation of electrostatic potential by successive addition of active site residues show that their roles on the reaction are similar in both classes despite the difference in the organization of the

  7. Derepression of Synthesis of the Aminoacyl-Transfer Ribonucleic Acid Synthetases for the Branched-Chain Amino Acids of Escherichia coli

    PubMed Central

    McGinnis, Etheleen; Williams, Ann C.; Williams, L. S.

    1974-01-01

    The kinetics of derepression of valyl-, isoleucyl-, and leucyl-transfer ribonucleic acid (tRNA) synthetase formation was examined during valine-, isoleucine-, and leucine-limited growth. When valine was limiting growth, valyl-tRNA synthetase formation was maximally derepressed within 5 min, whereas the rates of synthesis of isoleucyl-, and leucyl-tRNA synthetases were unchanged. Isoleucine-restricted growth caused a maximal derepression of isoleucyl-tRNA synthetase formation in 5 min and derepression of valyl-tRNA synthetase formation in 15 min with no effect on leucyl-tRNA synthetase formation. When leucine was limiting growth, leucyl-tRNA synthetase formation was immediately derepressed, whereas valyl- and isoleucyl-tRNA synthetase formation was unaffected by manipulation of the leucine supply to the cells. These results support our previous findings that valyl-tRNA synthetase formation is subject to multivalent repression control by both isoleucine and valine. In contrast, repression control of iso-leucyl- and leucyl-tRNA synthetase formation is specifically mediated by the supply of the cognate amino acid. PMID:4604302

  8. Mirror image alternative interaction patterns of the same tRNA with either class I arginyl-tRNA synthetase or class II aspartyl-tRNA synthetase.

    PubMed Central

    Sissler, M; Eriani, G; Martin, F; Giegé, R; Florentz, C

    1997-01-01

    Gene cloning, overproduction and an efficient purification protocol of yeast arginyl-tRNA synthetase (ArgRS) as well as the interaction patterns of this protein with cognate tRNAArgand non-cognate tRNAAspare described. This work was motivated by the fact that the in vitro transcript of tRNAAspis of dual aminoacylation specificity and is not only aspartylated but also efficiently arginylated. The crystal structure of the complex between class II aspartyl-tRNA synthetase (AspRS) and tRNAAsp, as well as early biochemical data, have shown that tRNAAspis recognized by its variable region side. Here we show by footprinting with enzymatic and chemical probes that transcribed tRNAAspis contacted by class I ArgRS along the opposite D arm side, as is homologous tRNAArg, but with idiosyncratic interaction patterns. Besides protection, footprints also show enhanced accessibility of the tRNAs to the structural probes, indicative of conformational changes in the complexed tRNAs. These different patterns are interpreted in relation to the alternative arginine identity sets found in the anticodon loops of tRNAArgand tRNAAsp. The mirror image alternative interaction patterns of unmodified tRNAAspwith either class I ArgRS or class II AspRS, accounting for the dual identity of this tRNA, are discussed in relation to the class defining features of the synthetases. This study indicates that complex formation between unmodified tRNAAspand either ArgRS and AspRS is solely governed by the proteins. PMID:9396794

  9. Heat welding of non-orthogonal X-junction of single-walled carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Yang, Xueming; Han, Zhonghe; Li, Yonghua; Chen, Dongci; Zhang, Pu; To, Albert C.

    2012-09-01

    Though X-junctions of single-walled carbon nanotubes (SWCNTs) have been intensively studied, studies concerning non-orthogonal X-junctions are still very rare. In this paper, the heat welding of defect-free non-orthogonal X-junctions with different crossed angles are investigated by molecular dynamics simulations. The difference between the heat welding of non-orthogonal and orthogonal X-junctions is described, and the angle effect on the configuration and stability of the heat welded non-orthogonal X-junctions is discussed. Compared with the orthogonal X-junction, two crossed SWCNTs with a smaller non-orthogonal angle are easier to join by heat welding, and this may be an important reason why the large tubes are difficult to join, whereas large nanotube bundles are easier to observe in experiments.

  10. Inhibitory effect of quinolone antimicrobial and nonsteroidal anti-inflammatory drugs on a medium chain acyl-CoA synthetase.

    PubMed

    Kasuya, F; Hiasa, M; Kawai, Y; Igarashi, K; Fukui, M

    2001-08-01

    The inhibitory effects of quinolone antimicrobial agents and nonsteroidal anti-inflammatory drugs on purified mouse liver mitochondrial medium chain acyl-CoA synthetase catalyzing the first reaction of glycine conjugation were examined, using hexanoic acid as a substrate. Enoxacin, ofloxacin, nalidixic acid, diflunisal, salicylic acid, 2-hydroxynaphthoic acid, and 2-hydroxydodecanoic acid, which do not act as substrates, were potent inhibitors. Diflunisal, nalidixic acid, salicylic acid, 2-hydroxynaphthoic acid, and 2-hydroxydodecanoic acid inhibited competitively this medium chain acyl-CoA synthetase with K(i) values of 0.6, 12.4, 19.6, 13.4, and 15.0 microM, respectively. Enoxacin and ofloxacin inhibited this medium chain acyl-CoA synthetase in a mixed-type manner with K(i) values of 23.7 and 38.2 microM, respectively. Felbinac, which is a substrate, inhibited the activity of this medium chain acyl-CoA synthetase for hexanoic acid (IC50 = 25 microM). The concomitant presence of enoxacin and felbinac strongly inhibited this medium chain acyl-CoA synthetase. These findings indicate that medium chain acyl-CoA synthetases may be influenced by quinolone antimicrobial and nonsteroidal anti-inflammatory drugs. PMID:11434910

  11. Expression of acyl-CoA synthetase 5 reflects the state of villus architecture in human small intestine.

    PubMed

    Gassler, Nikolaus; Kopitz, Jürgen; Tehrani, Arman; Ottenwälder, Birgit; Schnölzer, Martina; Kartenbeck, Jürgen; Lyer, Stefan; Autschbach, Frank; Poustka, Annemarie; Otto, Herwart F; Mollenhauer, Jan

    2004-02-01

    Several disorders of the small intestine are associated with disturbances in villus architecture. Thus, an understanding of the molecular mechanisms associated with the differentiation of villi represents an important step in the improvement of the understanding of small intestinal pathology. Screening of antibodies from a hybridoma library led to the identification of an acyl-CoA synthetase 5-specific monoclonal antibody. Protein synthesis, mRNA expression, and the enzyme activity of acyl-CoA synthetase 5 were studied by several methods in human small intestinal tissues with Crohn's disease or coeliac disease, respectively. Acyl-CoA synthetase 5 mRNA and protein levels were substantially reduced in injured small intestinal mucosa. Moreover, impaired synthesis of the acyl-CoA synthetase 5 protein was reflected by a decrease in intramucosal enzyme activity. Subtle changes of the acyl-CoA synthetase 5 pattern correlate with conversion of intestinal epithelial cells to a gastric phenotype. These results suggest that deranged acyl-CoA synthetase 5 expression, synthesis, and activity are closely related to the state of villus architecture and epithelial homeostasis in human small intestine. PMID:14743501

  12. Evidence that peroxisomal acyl-CoA synthetase is located at the cytoplasmic side of the peroxisomal membrane.

    PubMed Central

    Mannaerts, G P; Van Veldhoven, P; Van Broekhoven, A; Vandebroek, G; Debeer, L J

    1982-01-01

    1. Subfractionation by isopycnic density-gradient centrifugation in self-generating Percoll gradients of peroxisome-rich fractions prepared by differential centrifugation confirmed the presence of acyl-CoA synthetase in peroxisomes. Peroxisomes did not contain nicotinamide or adenine nucleotides other than CoA. 2. The gradient fractions most enriched in peroxisomes were pooled and the peroxisomes sedimented by centrifugation, resulting in a 50-fold-purified peroxisomal preparation as revealed by marker enzyme analysis. 3. Palmitate oxidation by intact purified peroxisomes was CoA-dependent, whereas palmitoyl-CoA oxidation was not, demonstrating that the peroxisomal CoA was available for the thiolase reaction, located in the peroxisomal matrix, but not for acyl-CoA synthetase. This suggests that the latter enzyme is located at the cytoplasmic side of the peroxisomal membrane. 4. Additional evidence for this location of peroxisomal acyl-CoA synthetase was as follows. Mechanical disruption of purified peroxisomes resulted in the release of catalase from the broken organelles, but not of acyl-CoA synthetase, indicating that the enzyme was membrane-bound. Acyl-CoA synthetase was not latent, despite the fact that at least one of its substrates appears to have a limited membrane permeability, as evidenced by the presence of CoA in purified peroxisomes. Finally, Pronase, a proteinase that does not penetrate the peroxisomal membrane, almost completely inactivated the acyl-CoA synthetase of intact peroxisomes. PMID:7115321

  13. 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

  14. 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.

  15. Inhibition of Plasmodium falciparum dihydropteroate synthetase and growth in vitro by sulfa drugs.

    PubMed Central

    Zhang, Y; Meshnick, S R

    1991-01-01

    The Michaelis-Menten inhibitory constants (Kis) and the concentrations required for 50% inhibition of the Plasmodium falciparum dihydropteroate synthetase were determined for six sulfa drugs. These drugs inhibited the in vitro growth of P. falciparum (50% lethal concentration) at concentrations of 30 to 500 nM; these concentrations were 100 to 1,000 times lower than the concentrations required for 50% inhibition and Kis (6 to 500 microM). The uptake of p-aminobenzoic acid was not inhibited by the sulfa drugs. However, infected erythrocytes took up more labeled sulfamethoxazole than did uninfected erythrocytes. Thus, the concentration of sulfa drugs by malaria parasites may explain how sulfa drugs inhibit in vitro growth of parasites through the inhibition of dihydropteroate synthetase. PMID:2024960

  16. Primary structure of the Saccharomyces cerevisiae gene for methionyl-tRNA synthetase.

    PubMed Central

    Walter, P; Gangloff, J; Bonnet, J; Boulanger, Y; Ebel, J P; Fasiolo, F

    1983-01-01

    The sequence of a 5-kilobase DNA insert containing the structural gene for yeast cytoplasmic methionyl-tRNA synthetase has been determined and a unique open reading frame of 2,253 nucleotides encoding a polypeptide chain of 751 amino acids (Mr, 85,500) has been characterized. The data obtained on the purified enzyme (subunit size, amino acid composition, and COOH-terminal sequence) are consistent with the gene structure. The protein sequence deduced from the nucleotide sequence reveals no obvious internal repeats. This protein sequence shows a high degree of homology with that of Escherichia coli methionyl-tRNA synthetase within a region that forms the putative methionyl adenylate binding site. This strongly suggests that both proteins derive from a common ancestor. PMID:6341994

  17. 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.

  18. 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.

  19. Structure of an unusual S-adenosylmethionine synthetase from Campylobacter jejuni.

    PubMed

    Zano, Stephen P; Pavlovsky, Alexander G; Viola, Ronald E

    2014-02-01

    S-Adenosylmethionine (AdoMet) participates in a wide range of methylation and other group-transfer reactions and also serves as the precursor for two groups of quorum-sensing molecules that function as regulators of the production of virulence factors in Gram-negative bacteria. The synthesis of AdoMet is catalyzed by AdoMet synthetases (MATs), a ubiquitous family of enzymes found in species ranging from microorganisms to mammals. The AdoMet synthetase from the bacterium Campylobacter jejuni (cjMAT) is an outlier among this homologous enzyme family, with lower sequence identity, numerous insertions and substitutions, and higher catalytic activity compared with other bacterial MATs. Alterations in the structure of this enzyme provide an explanation for its unusual dimeric quaternary structure relative to the other MATs. Taken together with several active-site substitutions, this new structure provides insights into its improved kinetic properties with alternative substrates. PMID:24531478

  20. Halofuginone and other febrifugine derivatives inhibit prolyl-tRNA synthetase

    PubMed Central

    Keller, Tracy L.; Zocco, Davide; Sundrud, Mark S.; Hendrick, Margaret; Edenius, Maja; Yum, Jina; Kim, Yeon-Jin; Lee, Hak-kyo; Cortese, Joseph F.; Wirth, Dyann; Dignam, John David; Rao, Anjana; Yeo, Chang-Yeol; Mazitschek, Ralph; Whitman, Malcolm

    2011-01-01

    Febrifugine, one of the fifty fundamental herbs of traditional Chinese medicine, has been characterized for its therapeutic activity whilst its molecular target has remained unknown. Febrifugine derivatives have been used to treat malaria, cancer, fibrosis, and inflammatory disease. We recently demonstrated that halofuginone (HF), a widely studied derivative of febrifugine, inhibits the development of Th17-driven autoimmunity in a mouse model of multiple sclerosis by activating the amino acid response pathway (AAR). Here we show that HF binds glutamyl-prolyl-tRNA synthetase (EPRS) inhibiting prolyl-tRNA synthetase activity; this inhibition is reversed by the addition of exogenous proline or EPRS. We further show that inhibition of EPRS underlies the broad bioactivities of this family of natural products. This work both explains the molecular mechanism of a promising family of therapeutics, and highlights the AAR pathway as an important drug target for promoting inflammatory resolution. PMID:22327401

  1. Escherichia coli proline tRNA: structure and recognition sites for prolyl-tRNA synthetase.

    PubMed

    Hasegawa, T; Yokogawa, T

    2000-01-01

    A major proline tRNA was purified from bulk Escherichia coli A19 tRNA by affinity chromatography with a biotinylated DNA probe. Its nucleotide sequence including modified nucleotides was determined by the post-labelling technique. In order to study the recognition sites of this proline tRNA for prolyl-tRNA synthetase, various mutant transcripts were prepared using an in vitro transcription system with T7 RNA polymerase. Based on the results of in vitro kinetic analyses of mutant transcripts, it was concluded that the second and third letters, G35 and G36, of the anticodon, G37 of the anticodon loop, the discriminator base A73, G72 of the acceptor stem, G49 and U17A that existed in the corner of an L-shaped structure are the recognition sites of proline tRNA for prolyl-tRNA synthetase. PMID:12903242

  2. Recoding Aminoacyl-tRNA Synthetases for Synthetic Biology by Rational Protein-RNA Engineering

    PubMed Central

    2015-01-01

    We have taken a rational approach to redesigning the amino acid binding and aminoacyl–tRNA pairing specificities of bacterial glutaminyl–tRNA synthetase. The four-stage engineering incorporates generalizable design principles and improves the pairing efficiency of noncognate glutamate with tRNAGln by over 105-fold compared to the wild-type enzyme. Better optimized designs of the protein–RNA complex include substantial reengineering of the globular core region of the tRNA, demonstrating a role for specific tRNA nucleotides in specifying the identity of the genetically encoded amino acid. Principles emerging from this engineering effort open new prospects for combining rational and genetic selection approaches to design novel aminoacyl–tRNA synthetases that ligate noncanonical amino acids onto tRNAs. This will facilitate reconstruction of the cellular translation apparatus for applications in synthetic biology. PMID:25310879

  3. Dual inhibitory effects of dimethyl sulfoxide on poly(ADP-ribose) synthetase.

    PubMed

    Banasik, M; Ueda, K

    1999-01-01

    Dimethyl sulfoxide (DMSO), a solvent popularly used for dissolving water-insoluble compounds, is a weak inhibitor of poly(ADP-ribose) synthetase, that is a nuclear enzyme producing (ADP-ribose)n from NAD+. The inhibitory mode and potency depend on the concentration of substrate, NAD+, as well as the temperature of the reaction; at micromolar concentrations of NAD+, the inhibition by DMSO is biphasic at 37 degrees C, but is monophasic and apparently competitive with NAD+ at 25 degrees C. DMSO, on the other hand, diminishes dose-dependently and markedly the inhibitory potency of benzamide and other inhibitors. Other organic solvents, ethanol and methanol, also show a biphasic effect on the synthetase activity at different concentrations. PMID:10445046

  4. S-adenosyl-L-methionine synthetase and phospholipid methyltransferase are inhibited in human cirrhosis.

    PubMed

    Duce, A M; Ortíz, P; Cabrero, C; Mato, J M

    1988-01-01

    We have measured the activity S-adenosyl-L-methionine synthetase in liver biopsies from a group of controls (n = 17) and in 26 cirrhotics (12 alcoholic and 14 posthepatic). The activity of this enzyme was markedly reduced in the group of cirrhotics (285 +/- 32 pmoles per min per mg protein) when compared with that observed in controls (505 +/- 37 pmoles per min per mg protein). No differences in S-adenosyl-L-methionine synthetase was observed between both groups of cirrhotics. Similarly, a marked reduction in the activity phospholipid methyltransferase was also observed in liver biopsies from the same group of cirrhotics (105 +/- 12 pmoles per min per mg protein) when compared with the control subjects (241 +/- 13 pmoles per min per mg protein). Again, no difference in the activity of this enzyme was observed between both groups of cirrhotics. These results indicated a marked deficiency in the metabolism of S-adenosyl-L-methionine in cirrhosis. PMID:3338721

  5. 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

  6. Effects of prostaglandins and prostaglandin synthetase inhibitors on acutely obstructed kidneys in the dog.

    PubMed

    Zwergel, U; Zwergel, T; Ziegler, M

    1991-01-01

    An intact canine model was developed to study the effects of prostaglandins (PG) and prostaglandin synthetase inhibitors on acutely obstructed kidneys. Totally implanted nephrostomy tubes were placed to measure renal pelvic pressure. Complete ureteral obstruction was obtained with a Fogarty balloon catheter inflated in the distal ureter; by this method renal pelvic pressure reached 40-50 mm Hg. Renal pelvic pressure was reduced after intravenous indomethacin and dipyrone administration, whereas blood pressure showed no major changes. Exogenous prostaglandins had both immediate and contrary effects: PGE2 caused a significant decrease, whereas PGF2 alpha caused a significant increase in renal pelvic and blood pressure. The reduced rise in renal pelvic pressure appears to be the main reason for the analgesic effects of prostaglandin synthetase inhibitors. The efficiency of these drugs in the treatment of renal colic is supported by this study, that of prostaglandins cannot be proved. PMID:1792708

  7. 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.

  8. 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.

  9. 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.

  10. Aspects of surface generation in orthogonal ultraprecision machining

    SciTech Connect

    Lucca, D.A.; Seo, Y.W.; Rhorer, R.L.

    1994-10-01

    The depth of the plastically deformed layer at the workpiece surface which resulted in the orthogonal ultraprecision machining of Cu over the range of uncut chip thicknesses of 0.01-10 {mu}m was investigated. Two tools with the same nominal geometry but with differing edge geometries were used to machine both Te-Cu and fine grain Cu. Tool edge geometries were characterized by atomic force microscopy, taking into account the AFM cantilever tip radius. Magnitudes of the measured depths appear to be consistent with values reported in the literature and those arrived at by simple analyses.

  11. Basis difference method for orthogonal systems on a surface

    NASA Astrophysics Data System (ADS)

    Korobitsyn, V. A.

    2011-07-01

    The basis operator method intended for constructing systems of difference approximations to differential operators in vector and tensor analysis is extended to orthogonal systems on a surface. A class of completely conservative differential-difference schemes for continuum mechanics in Lagrangian variables is constructed. Basis operators are constructed using the finite volume equation, consistency conditions for discrete operators of the first derivative, and consistent projection operators for grid functions. A system of differential-difference continuum mechanics equations on a surface is obtained, which implies all conservation laws typical of the continuum case, including additional ones. A stability estimate is derived for discrete equations of an incompressible viscous fluid.

  12. 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

  13. The Local Orthogonality Between Quantum States and Entanglement Decomposition

    NASA Astrophysics Data System (ADS)

    Kim, Sunho; Wu, Junde; Zhang, Lin; Cho, Minhyung

    2016-06-01

    In the paper, we show that when a quantum state can be decomposed as a convex combination of locally orthogonal mixed states, its entanglement can be decomposed into the entanglement of these mixed states without losing them. The obtained result generalizes a corresponding one proved by Horodecki (Acta Phys. Slov. 48, 141 1998). But, for the entanglement cost it requires certain conditions for holding the decomposition, and the distillable entanglement only has a week result as inequality. Finally, we presented an example to show that the conditions of our conclusions are existence.

  14. Recurrence relations of the multi-indexed orthogonal polynomials. III

    NASA Astrophysics Data System (ADS)

    Odake, Satoru

    2016-02-01

    In Paper II, we presented conjectures of the recurrence relations with constant coefficients for the multi-indexed orthogonal polynomials of Laguerre, Jacobi, Wilson, and Askey-Wilson types. In this paper we present a proof for the Laguerre and Jacobi cases. Their bispectral properties are also discussed, which gives a method to obtain the coefficients of the recurrence relations explicitly. This paper extends to the Laguerre and Jacobi cases the bispectral techniques recently introduced by Gómez-Ullate et al. [J. Approx. Theory 204, 1 (2016); e-print arXiv:1506.03651 [math.CA

  15. 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.

  16. Conditioning analysis of incomplete Cholesky factorizations with orthogonal dropping

    SciTech Connect

    Napov, Artem

    2012-03-16

    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.

  17. Orthogonal cutting characteristics of graphite/epoxy composite material

    NASA Astrophysics Data System (ADS)

    Wang, D. H.; Ramulu, M.; Wern, C. W.

    Orthogonal cutting experimental study was conducted to investigate the machining characteristics of Graphite/Epoxy (Gr/Ep) composite material. Machining characteristics were evaluated in terms of cutting forces, chip formation, and surface morphology of unidirectional Gr/Ep composite material of different fiber orientations. The cutting forces were measured by a three-dimensional circular-type dynamometer. Chips were examined under Scanning Electron Microscopy (SEM) and the machined surface morphology was investigated by measuring the surface roughness and by observing SEM photographs. Cutting forces, chip formation process and the surface morphology of machined surface were found to be highly dependent on the fiber orientations with respect to the cutting direction.

  18. 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).

  19. Spin Symmetry and Size Consistency of Strongly Orthogonal Geminals.

    PubMed

    Jeszenszki, Péter; Surján, Péter R; Szabados, Ágnes

    2015-07-14

    An overview of geminal-based wavefunctions is given, allowing for singlet-triplet mixing within the two-electron units. Spin contamination of the total wavefunction (obtained as an antisymmetrized product) is restored by spin projection. Full variation after projection is examined for two models. One is the long known spin-projected, extended Hartree-Fock (EHF). The other is a yet unexplored function, termed spin-projected, extended antisymmetrized product of strongly orthogonal geminals (EAPSG). Studies on size consistency are presented for both models. Numerical evaluation of EHF and EAPSG is performed for small test systems (H4 and H8). PMID:26575746

  20. Orthogonal linear group-subgroup pairs with the same invariants

    NASA Astrophysics Data System (ADS)

    Solomon, S.

    2005-03-01

    The main theorem of Galois theory states that there are no finite group-subgroup pairs with the same invariants. On the other hand, if we consider complex linear reductive groups instead of finite groups, the analogous statement is no longer true: There exist counterexample group-subgroup pairs with the same invariants. However, it's possible to classify all these counterexamples for certain types of groups. In [16], we provided the classification for connected complex irreducible groups, and, in this paper, for connected complex orthogonal groups, i.e., groups that preserve some non-degenerate quadratic form.

  1. Universality of Mesoscopic Fluctuations for Orthogonal Polynomial Ensembles

    NASA Astrophysics Data System (ADS)

    Breuer, Jonathan; Duits, Maurice

    2016-03-01

    We prove that the fluctuations of mesoscopic linear statistics for orthogonal polynomial ensembles are universal in the sense that two measures with asymptotic recurrence coefficients have the same asymptotic mesoscopic fluctuations (under an additional assumption on the local regularity of one of the measures). The convergence rate of the recurrence coefficients determines the range of scales on which the limiting fluctuations are identical. Our main tool is an analysis of the Green's function for the associated Jacobi matrices.As a particular consequencewe obtain a central limit theorem for the modified Jacobi Unitary Ensembles on all mesoscopic scales.

  2. Proton lifetime in orthogonal theories of family unification

    NASA Astrophysics Data System (ADS)

    Bagger, Jonathan; Dimopoulos, Savas; Massó, Eduard

    1984-09-01

    We compute the proton lifetime in recently proposed orthogonal theories of family unification. For ΛMS = 100 MeV, we find a partial lifetime τ(p-->e+π0) of 5.9×1031+/-1 yr, where the error in the exponent comes from uncertainties in the hadronic wave function. Important decay products include electrons, neutrinos and non-strange mesons. Fulbright Fellow. On leave of absence from Departament de Física Teòrica, Universitat Autónoma de Barcelona, Belaterra, Spain.

  3. 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.

  4. Inhibition of isoleucyl-transfer ribonucleic acid synthetase in Echerichia coli by pseudomonic acid

    PubMed Central

    Hughes, Julia; Mellows, Graham

    1978-01-01

    The mode of action of the antibiotic pseudomonic acid has been studied in Escherichia coli. Pseudomonic acid strongly inhibits protein and RNA synthesis in vivo. The antibiotic had no effect on highly purified DNA-dependent RNA polymerase and showed only a weak inhibitory effect on a poly(U)-directed polyphenylalanine-forming ribosomal preparation. Chloramphenicol reversed inhibition of RNA synthesis in vivo. Pseudomonic acid had little effect on RNA synthesis in a regulatory mutant, E. coli B AS19 RCrel, whereas protein synthesis was strongly inhibited. In pseudomonic acid-treated cells, increased concentrations of ppGpp, pppGpp and ATP were observed, but the GTP pool size decreased, suggesting that inhibition of RNA synthesis is a consequence of the stringent control mechanism imposed by pseudomonic acid-induced deprivation of an amino acid. Of the 20 common amino acids, only isoleucine reversed the inhibitory effect in vivo. The antibiotic was found to be a powerful inhibitor of isoleucyl-tRNA synthetase both in vivo and in vitro. Of seven other tRNA synthetases assayed, only a weak inhibitory effect on phenylalanyl-tRNA synthetase was observed; this presumably accounted for the weak effect on polyphenylalanine formation in a ribosomal preparation. Pseudomonic acid also significantly de-repressed threonine deaminase and transaminase B activity, but not dihydroxyacid dehydratase (isoleucine-biosynthetic enzymes) by decreasing the supply of aminoacylated tRNAIle. Pseudomonic acid is the second naturally occurring inhibitor of bacterial isoleucyl-tRNA synthetase to be discovered, furanomycin being the first. PMID:365175

  5. Glutamine synthetase gene expression during the regeneration of the annelid Enchytraeus japonensis.

    PubMed

    Niva, Cintia Carla; Lee, Jae Min; Myohara, Maroko

    2008-01-01

    Enchytraeus japonensis is a highly regenerative oligochaete annelid that can regenerate a complete individual from a small body fragment in 4-5 days. In our previous study, we performed complementary deoxyribonucleic acid subtraction cloning to isolate genes that are upregulated during E. japonensis regeneration and identified glutamine synthetase (gs) as one of the most abundantly expressed genes during this process. In the present study, we show that the full-length sequence of E. japonensis glutamine synthetase (EjGS), which is the first reported annelid glutamine synthetase, is highly similar to other known class II glutamine synthetases. EjGS shows a 61-71% overall amino acid sequence identity with its counterparts in various other animal species, including Drosophila and mouse. We performed detailed expression analysis by in situ hybridization and reveal that strong gs expression occurs in the blastemal regions of regenerating E. japonensis soon after amputation. gs expression was detectable at the cell layer covering the wound and was found to persist in the epidermal cells during the formation and elongation of the blastema. Furthermore, in the elongated blastema, gs expression was detectable also in the presumptive regions of the brain, ventral nerve cord, and stomodeum. In the fully formed intact head, gs expression was also evident in the prostomium, brain, the anterior end of the ventral nerve cord, the epithelium of buccal and pharyngeal cavities, the pharyngeal pad, and in the esophageal appendages. In intact E. japonensis tails, gs expression was found in the growth zone in actively growing worms but not in full-grown individuals. In the nonblastemal regions of regenerating fragments and in intact worms, gs expression was also detected in the nephridia, chloragocytes, gut epithelium, epidermis, spermatids, and oocytes. These results suggest that EjGS may play roles in regeneration, nerve function, cell proliferation, nitrogenous waste excretion

  6. 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.

  7. 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.

  8. A broadly applicable continuous spectrophotometric assay for measuring aminoacyl-tRNA synthetase activity.

    PubMed Central

    Lloyd, A J; Thomann, H U; Ibba, M; Söll, D

    1995-01-01

    We describe a convenient, simple and novel continuous spectrophotometric method for the determination of aminoacyl-tRNA synthetase activity. The assay relies upon the measurement of inorganic pyrophosphate generated in the first step of the aminoacylation of a tRNA. Pyrophosphate release is coupled to inorganic pyrophosphatase, to generate phosphate, which in turn is used as the substrate of purine nucleoside phosphorylase to catalyze the N-glycosidic cleavage of 2-amino 6-mercapto 7-methylpurine ribonucleoside. Of the reaction products, ribose 1-phosphate and 2-amino 6-mercapto 7-methylpurine, the latter has a high absorbance at 360 nm relative to the nucleoside and hence provides a spectrophotometric signal that can be continuously followed. The non-destructive nature of the spectrophotometric assay allowed the re-use of the tRNAs in question in successive experiments. The usefulness of this method was demonstrated for glutaminyl-tRNA synthetase (GlnRS) and tryptophanyl-tRNA synthetase. Initial velocities measured using this assay correlate closely with those assayed by quantitation of [3H]Gln-tRNA or [14C]Trp-tRNA formation respectively. In both cases amino acid transfer from the aminoacyl adenylate to the tRNA represents the rate determining step. In addition, aminoacyl adenylate formation by aspartyl-tRNA synthetase was followed and provided a more sensitive means of active site titration than existing techniques. Finally, this novel method was used to provide direct evidence for the cooperativity of tRNA and ATP binding to GlnRS. PMID:7659511

  9. 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

  10. Structure and regulation of expression of the Bacillus subtilis valyl-tRNA synthetase gene.

    PubMed Central

    Luo, D; Leautey, J; Grunberg-Manago, M; Putzer, H

    1997-01-01

    We have sequenced the valyl-tRNA synthetase gene (valS) of Bacillus subtilis and found an open reading frame coding for a protein of 880 amino acids with a molar mass of 101,749. The predicted amino acid sequence shares strong similarity with the valyl-tRNA synthetases from Bacillus stearothermophilus, Lactobacillus casei, and Escherichia coli. Extracts of B. subtilis strains overexpressing the valS gene on a plasmid have increased valyl-tRNA aminoacylation activity. Northern analysis shows that valS is cotranscribed with the folC gene (encoding folyl-polyglutamate synthetase) lying downstream. The 300-bp 5' noncoding region of the gene contains the characteristic regulatory elements, T box, "specifier codon" (GUC), and rho-independant transcription terminator of a gene family in gram-positive bacteria that encodes many aminoacyl-tRNA synthetases and some amino acid biosynthetic enzymes and that is regulated by tRNA-mediated antitermination. We have shown that valS expression is induced by valine limitation and that the specificity of induction can be switched to threonine by changing the GUC (Val) specifier triplet to ACC (Thr). Overexpression of valS from a recombinant plasmid leads to autorepression of a valS-lacZ transcriptional fusion. Like induction by valine starvation, autoregulation of valS depends on the presence of the GUC specifier codon. Disruption of the valS gene was not lethal, suggesting the existence of a second gene, as is the case for both the thrS and the tyrS genes. PMID:9098041

  11. Virtual Screening to Identify Lead Inhibitors for Bacterial NAD Synthetase (NADs)

    PubMed Central

    Moro, Whitney Beysselance; Yang, Zhengrong; Kane, Tasha; Brouillette, Christie G.; Brouillette, Wayne J.

    2009-01-01

    Virtual screening was employed to identify new drug-like inhibitors of NAD synthetase (NADs) as antibacterial agents. Four databases of commercially available compounds were docked against three subsites of the NADs active site using FlexX in conjunction with CScore. Over 200 commercial compounds were purchased and evaluated in enzyme inhibition and antibacterial assays. 18 compounds inhibited NADs at or below 100 μM (7.6% hit rate), and two were selected for future SAR studies. PMID:19249205

  12. Effect of post-silking drought on nitrogen partitioning and gene expression patterns of glutamine synthetase and asparagine synthetase in two maize (Zea mays L.) varieties.

    PubMed

    Li, Yajun; Wang, Meiling; Zhang, Fengxia; Xu, Yadong; Chen, Xiaohong; Qin, Xiaoliang; Wen, Xiaoxia

    2016-05-01

    Glutamine synthetase (GS) and asparagine synthetase (AS) are proposed to have important function in plant nitrogen (N) remobilization, but their roles under drought stress are not well defined. In this study, the expression dynamics of GS and AS genes were analyzed in two maize varieties (ZD958 and NH101) in relation to post-silking drought stress induced nitrogen partitioning. ZD958 was a 'stay-green' variety with 5% nitrogen harvest index (NHI) lower than NH101. From silking to maturity, the amount of nitrogen remobilized from ear-leaves in ZD958 was evidently lower than NH101, and post-silking drought stress increased the nitrogen remobilization for both varieties. In ear-leaves, the expression of ZmGln1-3 was enhanced under drought stress. Three AS genes (ZmAS1, ZmAS2 and ZmAS3) were differentially regulated by post-silking drought treatment, of which the expression of ZmAS3 was stimulated at late stage of leaf senescence. In NH101, the expression level of ZmAS3 was markedly higher than that in ZD958. In developing grains, there were no significant differences in expression patterns of GS and AS genes between well water and drought treated plants. Drought stress altered maize N partitioning at the whole-plant level, and the up-regulation of GS and AS genes may contribute to the higher leaf nitrogen remobilization when exposed to drought treatments. PMID:26913793

  13. Dihydroaeruginoic acid synthetase and pyochelin synthetase, products of the pchEF genes, are induced by extracellular pyochelin in Pseudomonas aeruginosa.

    PubMed

    Reimmann, C; Serino, L; Beyeler, M; Haas, D

    1998-11-01

    The siderophore pyochelin of Pseudomonas aeruginosa is derived from one molecule of salicylate and two molecules of cysteine. Two cotranscribed genes, pchEF, encoding peptide synthetases have been identified and characterized. pchE was required for the conversion of salicylate to dihydroaeruginoate (Dha), the condensation product of salicylate and one cysteine residue and pchF was essential for the synthesis of pyochelin from Dha. The deduced PchE (156 kDa) and PchF (197 kDa) proteins had adenylation, thiolation and condensation/cyclization motifs arranged as modules which are typical of those peptide synthetases forming thiazoline rings. The pchEF genes were coregulated with the pchDCBA operon, which provides enzymes for the synthesis (PchBA) and activation (PchD) of salicylate as well as a putative thioesterase (PchC). Expression of a translational pchE'-'lacZ fusion was strictly dependent on the PchR regulator and was induced by extracellular pyochelin, the end product of the pathway. Iron replete conditions led to Fur (ferric uptake regulator)-dependent repression of the pchE'-'lacZ fusion. A translational pchD'-'lacZ fusion was also positively regulated by PchR and pyochelin and repressed by Fur and iron. Thus, autoinduction by pyochelin (or ferric pyochelin) and repression by iron ensure a sensitive control of the pyochelin pathway in P. aeruginosa. PMID:9846750

  14. Structural studies of lysyl-tRNA synthetase: conformational changes induced by substrate binding.

    PubMed

    Onesti, S; Desogus, G; Brevet, A; Chen, J; Plateau, P; Blanquet, S; Brick, P

    2000-10-24

    Lysyl-tRNA synthetase is a member of the class II aminoacyl-tRNA synthetases and catalyses the specific aminoacylation of tRNA(Lys). The crystal structure of the constitutive lysyl-tRNA synthetase (LysS) from Escherichia coli has been determined to 2.7 A resolution in the unliganded form and in a complex with the lysine substrate. A comparison between the unliganded and lysine-bound structures reveals major conformational changes upon lysine binding. The lysine substrate is involved in a network of hydrogen bonds. Two of these interactions, one between the alpha-amino group and the carbonyl oxygen of Gly 216 and the other between the carboxylate group and the side chain of Arg 262, trigger a subtle and complicated reorganization of the active site, involving the ordering of two loops (residues 215-217 and 444-455), a change in conformation of residues 393-409, and a rotation of a 4-helix bundle domain (located between motif 2 and 3) by 10 degrees. The result of these changes is a closing up of the active site upon lysine binding. PMID:11041850

  15. 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

  16. Resected RNA pseudoknots and their recognition by histidyl-tRNA synthetase

    PubMed Central

    Felden, Brice; Giegé, Richard

    1998-01-01

    Duplexes constituted by closed or open RNA circles paired to single-stranded oligonucleotides terminating with 3′-CCAOH form resected pseudoknots that are substrates of yeast histidyl-tRNA synthetase. Design of this RNA fold is linked to the mimicry of the pseudoknotted amino acid accepting branch of the tRNA-like domain from brome mosaic virus, known to be charged by tyrosyl-tRNA synthetases, with RNA minihelices recapitulating accepting branches of canonical tRNAs. Prediction of the histidylation function of the new family of minimalist tRNA-like structures relates to the geometry of resected pseudoknots that allows proper presentation to histidyl-tRNA synthetase of analogues of the histidine identity determinants N-1 and N73 present in tRNAs. This geometry is such that the analogue of the major N-1 histidine determinant in the RNA circles faces the analogue of the discriminator N73 nucleotide in the accepting oligonucleotides. The combination of identity elements found in tRNAHis species from archaea, eubacteria, and organelles (G-1/C73) is the most efficient for determining histidylation of the duplexes. The inverse combination (C-1/G73) leads to the worst histidine acceptors with charging efficiencies reduced by 2–3 orders of magnitude. Altogether, these findings open new perspectives for understanding evolution of tRNA identity and serendipitous RNA functions. PMID:9724720

  17. Resected RNA pseudoknots and their recognition by histidyl-tRNA synthetase.

    PubMed

    Felden, B; Giegé, R

    1998-09-01

    Duplexes constituted by closed or open RNA circles paired to single-stranded oligonucleotides terminating with 3'-CCAOH form resected pseudoknots that are substrates of yeast histidyl-tRNA synthetase. Design of this RNA fold is linked to the mimicry of the pseudoknotted amino acid accepting branch of the tRNA-like domain from brome mosaic virus, known to be charged by tyrosyl-tRNA synthetases, with RNA minihelices recapitulating accepting branches of canonical tRNAs. Prediction of the histidylation function of the new family of minimalist tRNA-like structures relates to the geometry of resected pseudoknots that allows proper presentation to histidyl-tRNA synthetase of analogues of the histidine identity determinants N-1 and N73 present in tRNAs. This geometry is such that the analogue of the major N-1 histidine determinant in the RNA circles faces the analogue of the discriminator N73 nucleotide in the accepting oligonucleotides. The combination of identity elements found in tRNAHis species from archaea, eubacteria, and organelles (G-1/C73) is the most efficient for determining histidylation of the duplexes. The inverse combination (C-1/G73) leads to the worst histidine acceptors with charging efficiencies reduced by 2-3 orders of magnitude. Altogether, these findings open new perspectives for understanding evolution of tRNA identity and serendipitous RNA functions. PMID:9724720

  18. Isolation and characterization of the gene coding for Escherichia coli arginyl-tRNA synthetase.

    PubMed Central

    Eriani, G; Dirheimer, G; Gangloff, J

    1989-01-01

    The gene coding for Escherichia coli arginyl-tRNA synthetase (argS) was isolated as a fragment of 2.4 kb after analysis and subcloning of recombinant plasmids from the Clarke and Carbon library. The clone bearing the gene overproduces arginyl-tRNA synthetase by a factor 100. This means that the enzyme represents more than 20% of the cellular total protein content. Sequencing revealed that the fragment contains a unique open reading frame of 1734 bp flanked at its 5' and 3' ends respectively by 247 bp and 397 bp. The length of the corresponding protein (577 aa) is well consistent with earlier Mr determination (about 70 kd). Primer extension analysis of the ArgRS mRNA by reverse transcriptase, located its 5' end respectively at 8 and 30 nucleotides downstream of a TATA and a TTGAC like element (CTGAC) and 60 nucleotides upstream of the unusual translation initiation codon GUG; nuclease S1 analysis located the 3'-end at 48 bp downstream of the translation termination codon. argS has a codon usage pattern typical for highly expressed E. coli genes. With the exception of the presence of a HVGH sequence similar to the HIGH consensus element, ArgRS has no relevant sequence homologies with other aminoacyl-tRNA synthetases. Images PMID:2668891

  19. Crystal structure of histidyl-tRNA synthetase from Escherichia coli complexed with histidyl-adenylate.

    PubMed Central

    Arnez, J G; Harris, D C; Mitschler, A; Rees, B; Francklyn, C S; Moras, D

    1995-01-01

    The crystal structure at 2.6 A of the histidyl-tRNA synthetase from Escherichia coli complexed with histidyl-adenylate has been determined. The enzyme is a homodimer with a molecular weight of 94 kDa and belongs to the class II of aminoacyl-tRNA synthetases (aaRS). The asymmetric unit is composed of two homodimers. Each monomer consists of two domains. The N-terminal catalytic core domain contains a six-stranded antiparallel beta-sheet sitting on two alpha-helices, which can be superposed with the catalytic domains of yeast AspRS, and GlyRS and SerRS from Thermus thermophilus with a root-mean-square difference on the C alpha atoms of 1.7-1.9 A. The active sites of all four monomers are occupied by histidyl-adenylate, which apparently forms during crystallization. The 100 residue C-terminal alpha/beta domain resembles half of a beta-barrel, and provides an independent domain oriented to contact the anticodon stem and part of the anticodon loop of tRNA(His). The modular domain organization of histidyl-tRNA synthetase reiterates a repeated theme in aaRS, and its structure should provide insight into the ability of certain aaRS to aminoacylate minihelices and other non-tRNA molecules. Images PMID:7556055

  20. Alternative splicing creates two new architectures for human tyrosyl-tRNA synthetase

    PubMed Central

    Wei, Zhiyi; Xu, Zhiwen; Liu, Xiaotian; Lo, Wing-Sze; Ye, Fei; Lau, Ching-Fun; Wang, Feng; Zhou, Jie J.; Nangle, Leslie A.; Yang, Xiang-Lei; Zhang, Mingjie; Schimmel, Paul

    2016-01-01

    Many human tRNA synthetases evolved alternative functions outside of protein synthesis. These functions are associated with over 200 splice variants (SVs), most of which are catalytic nulls that engender new biology. While known to regulate non-translational activities, little is known about structures resulting from natural internal ablations of any protein. Here, we report analysis of two closely related, internally deleted, SVs of homodimeric human tyrosyl-tRNA synthetase (TyrRS). In spite of both variants ablating a portion of the catalytic core and dimer-interface contacts of native TyrRS, each folded into a distinct stable structure. Biochemical and nuclear magnetic resonance (NMR) analysis showed that the internal deletion of TyrRSΔE2–4 SV gave an alternative, neomorphic dimer interface ‘orthogonal’ to that of native TyrRS. In contrast, the internal C-terminal splice site of TyrRSΔE2–3 prevented either dimerization interface from forming, and yielded a predominantly monomeric protein. Unlike ubiquitous TyrRS, the neomorphs showed clear tissue preferences, which were distinct from each other. The results demonstrate a sophisticated structural plasticity of a human tRNA synthetase for architectural reorganizations that are preferentially elicited in specific tissues. PMID:26773056

  1. Localization of the L-glutamine synthetase gene to chromosome 1q23.

    PubMed

    Clancy, K P; Berger, R; Cox, M; Bleskan, J; Walton, K A; Hart, I; Patterson, D

    1996-12-15

    Glutamine synthetase (E.C. 6.3.1.2) is expressed throughout the body and plays an important role in controlling body pH and in removing ammonia from the circulation. The enzyme clears L-glutamate, the major neurotransmitter in the central nervous system, from neuronal synapses. The enzyme is a very sensitive marker of many disease and aging processes, especially those involving reactive oxygen species. This report describes the localization of the enzyme to chromosome 1 by PCR analysis of a human/rodent somatic cell hybrid panel. We also describe the localization of a recently described pseudogene to chromosome 9. Further localization of the glutamine synthetase gene locus to 1q23 was accomplished by fluorescence in situ hybridization. The glutamine synthetase gene was mapped to five CEPH megaYACs between the polymorphic PCR markers D1S117 and D1S466 by analysis of the Whitehead Institute's recently described chromosome 1 contig map. PMID:8975719

  2. Purification and characterization of fatty acyl-acyl carrier protein synthetase from Vibrio harveyi.

    PubMed Central

    Fice, D; Shen, Z; Byers, D M

    1993-01-01

    A Vibrio harveyi enzyme which catalyzes the ATP-dependent ligation of fatty acids to acyl carrier protein (ACP) has been purified 6,000-fold to apparent homogeneity by anion-exchange, gel filtration, and ACP-Sepharose affinity chromatography. Purified acyl-ACP synthetase migrated as a single 62-kDa band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and as an 80-kDa protein by gel filtration under reducing conditions. Activity of the purified enzyme was lost within hours in the absence of glycerol and low concentrations of Triton X-100. Acyl-ACP synthetase exhibited Kms for myristic acid, ACP, and ATP of 7 microM, 18 microM, and 0.3 mM, respectively. The enzyme was specific for adenine-containing nucleotides, and AMP was the product of the reaction. No covalent acyl-enzyme intermediate was observed. Enzyme activity was stimulated up to 50% by iodoacetamide but inhibited > 80% by N-ethylmaleimide: inhibition by the latter was prevented by ATP and ACP but not myristic acid. Dithiothreitol and sulfhydryl-directed reagents also influenced enzyme size, activity, and elution pattern on anion-exchange resins. The function of acyl-ACP synthetase has not been established, but it may be related to the capacity of V. harveyi to elongate exogenous fatty acids by an ACP-dependent mechanism. Images PMID:8384617

  3. The structure of tryptophanyl-tRNA synthetase from Giardia lamblia reveals divergence from eukaryotic homologs

    PubMed Central

    Arakaki, Tracy L; Carter, Megan; Napuli, Alberto J; Verlinde, Christophe L M J; Fan, Erkang; Zucker, Frank; Buckner, Frederick S; Van Voorhis, Wesley C; Hol, Wim G J; Merritt, Ethan A

    2010-01-01

    The 2.1 Å crystal structure of tryptophanyl-tRNA synthetase (TrpRS) from the diplomonad Giardia lamblia reveals that the N-terminus of this class I aminoacyl-tRNA synthetase forms a 16-residue α-helix. This helix replaces a β-hairpin that is required by human TrpRS for normal activity and has been inferred to play a similar role in all eukaryotic TrpRS. The primary sequences of TrpRS homologs from several basal eukaryotes including Giardia lack a set of three residues observed to stabilize interactions with this β-hairpin in the human TrpRS. Thus the present structure suggests that the activation reaction mechanism of TrpRS from the basal eukaryote G. lamblia differs from that of higher eukaryotes. Furthermore, the protein as observed in the crystal forms an (α2)2 homotetramer. The canonical dimer interface observed in all previous structures of tryptophanyl-tRNA synthetases is maintained, but in addition each N-terminal α-helix reciprocally interlocks with the equivalent helix from a second dimer to form a dimer of dimers. Although we have no evidence for tetramer formation in vivo, modeling indicates that the crystallographically observed tetrameric structure would be compatible with the tRNA binding mode used by dimeric TrpRS and TyrRS. PMID:20438846

  4. Mutants of Salmonella typhimurium with an Altered Leucyl-Transfer Ribonucleic Acid Synthetase

    PubMed Central

    Alexander, Renee R.; Calvo, J. M.; Freundlich, M.

    1971-01-01

    Two trifluoroleucine-resistant mutants of Salmonella typhimurium, strains CV69 and CV117, had an altered leucyl-transfer ribonucleic acid (tRNA) synthetase. The mutant enzymes had higher apparent Km values for leucine (ca. 10-fold) and lower specific activities (ca. twofold) than the parent enzyme when tested in crude extracts. Preparations of synthetase purified ca. 60-fold from the parent and strain CV117 differed sixfold in their leucine Km values. In addition, the mutant enzyme was inactivated faster than the parent enzyme at 50 C. The growth rates of strains CV69 and CV117 at 37 C were not significantly different from that of the parent, whereas at 42 C strain CV69 grew more slowly than the parent. Leucine-, valine-, and isoleucine-forming enzymes were partially derepressed when the mutants were grown in minimal medium; the addition of leucine repressed these enzymes to wild-type levels. During growth in minimal medium, the proportion of leucine tRNA that was charged in the mutants was about 75% of that in the parent. The properties of strain CV117 were shown to result from a single mutation located near gal at minute 18 on the genetic map. These studies suggest that leucyl-tRNA synthetase is involved in repression of the enzymes required for the synthesis of branched-chain amino acids. PMID:4928008

  5. Cloning and nucleotide sequence of the leucyl-tRNA synthetase gene of Bacillus subtilis.

    PubMed Central

    Vander Horn, P B; Zahler, S A

    1992-01-01

    The leucyl-tRNA synthetase gene (leuS) of Bacillus subtilis was cloned and sequenced. A mutation in the gene, leuS1, increases the transcription and expression of the ilv-leu operion, permitting monitoring of leuS alleles. The leuS1 mutation was mapped to 270 degrees on the chromosome. Sequence analysis showed that the mutation is a single-base substitution, possibly in a monocistronic operon. The leader mRNA predicted by the sequence would contain a number of possible secondary structures and a T box, a sequence observed upstream of leader mRNA terminators of Bacillus tRNA synthetases and the B. subtilis ilv-leu operon. The DNA of the B. subtilis leuS open reading frame is 48% identical to the leuS gene of Escherichia coli and is predicted to encode a polypeptide with 46% identity to the leucyl-tRNA synthetase of E. coli. PMID:1317842

  6. (p)ppGpp synthetases regulate the pathogenesis of zoonotic Streptococcus suis.

    PubMed

    Zhu, Jiawen; Zhang, Tengfei; Su, Zhipeng; Li, Lu; Wang, Dong; Xiao, Ran; Teng, Muye; Tan, Meifang; Zhou, Rui

    2016-10-01

    (p)ppGpp-mediated stringent response is one of the main adaption mechanism in bacteria, and the ability to adapt to environment is linked to the pathogenesis of bacterial pathogens. In the zoonotic pathogen Streptococcus suis, there are two (p)ppGpp synthetases, RelA and RelQ. To investigate the regulatory functions of (p)ppGpp/(p)ppGpp synthetases on the pathogenesis of S. suis, the phenotypes of the [(p)ppGpp(0)] mutant ΔrelAΔrelQ and its parental strain were compared. Light and electron microscopy observation showed that the mutant strain had a longer chain-length than its parental strain. Disruption of relA and relQ led to decreased adhesive and invasive ability to HEp-2 cells, and increased sensitivity to the blood killing and phagocytosis. Mouse infection experiments showed that the mutant strain was attenuated and easier to be cleaned up in vivo. Quantitative reverse transcription PCR (qRT-PCR) analysis indicated that the expressions of virulence related genes involving in morphology and virulence were down-regulated in the mutant strain. Our study demonstrated that the (p)ppGpp synthetases or (p)ppGpp can regulate the pathogenesis of this important zoonotic pathogen. PMID:27524648

  7. 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

  8. CphA2 is a novel type of cyanophycin synthetase in N2-fixing cyanobacteria.

    PubMed

    Klemke, Friederike; Nürnberg, Dennis J; Ziegler, Karl; Beyer, Gabriele; Kahmann, Uwe; Lockau, Wolfgang; Volkmer, Thomas

    2016-03-01

    Most cyanobacteria use a single type of cyanophycin synthetase, CphA1, to synthesize the nitrogen-rich polymer cyanophycin. The genomes of many N2-fixing cyanobacteria contain an additional gene that encodes a second type of cyanophycin synthetase, CphA2. The potential function of this enzyme has been debated due to its reduced size and the lack of one of the two ATP-binding sites that are present in CphA1. Here, we analysed CphA2 from Anabaena variabilis ATCC 29413 and Cyanothece sp. PCC 7425. We found that CphA2 polymerized the dipeptide β-aspartyl-arginine to form cyanophycin. Thus, CphA2 represents a novel type of cyanophycin synthetase. A cphA2 disruption mutant of A. variabilis was generated. Growth of this mutant was impaired under high-light conditions and nitrogen deprivation, suggesting that CphA2 plays an important role in nitrogen metabolism under N2-fixing conditions. Electron micrographs revealed that the mutant had fewer cyanophycin granules, but no alteration in the distribution of granules in its cells was observed. Localization of CphA2 by immunogold electron microscopy demonstrated that the enzyme is attached to cyanophycin granules. Expression of CphA1 and CphA2 was examined in Anabaena WT and cphA mutant cells. Whilst the CphA1 level increased upon nitrogen deprivation, the CphA2 level remained nearly constant. PMID:26781249

  9. Differential inactivation of alfalfa nodule glutamine synthetases by tabtoxinine-. beta. -lactam. [Pseudomonas syringae

    SciTech Connect

    Knight, T.J.; Unkefer, P.J.

    1987-04-01

    The presence of the pathogen Pseudomonas syringae pv. tabaci within the rhizosphere of nodulated alfalfa plants results in an increase in N/sub 2/-fixation potential and growth, but a 40-50% decrease in nodule glutamine synthetase (GS) activity, as compared to nodulated control plants. Tabtoxinine-..beta..-Lactam an exocellular toxin produced by Pseudomonas syringae pv tabaci irreversibly inhibits glutamine synthetase. Partial purification of nodule GS by DEAE-cellulose chromatography reveals two enzyme forms are present (GS/sub n1/ and GS/sub n2/). In vitro inactivation of the two glutamine synthetases associated with the nodule indicates a differential sensitivity to T-..beta..-L. The nodule specific GS/sub n1/ is much less sensitive to T-..beta..-L than the GS/sub n2/ enzyme, which was found to coelute with the root enzyme (GS/sub r/). However, both GS/sub n1/ and GS/sub n2/ are rapidly inactivated by methionine sulfoximine, another irreversible inhibitor of GS.

  10. Examination of the mechanism of sucrose synthetase by positional isotope exchange

    SciTech Connect

    Singh, A.N.; Hester, L.S.; Raushel, F.M.

    1987-02-25

    The mechanism of the sucrose synthetase reaction has been probed by the technique of positional isotope exchange. (beta-/sup 18/O/sub 2/, alpha beta-/sup 18/O)UDP-Glc has been synthesized starting from oxygen-18-labeled phosphate and the combined activities of carbamate kinase, hexokinase, phosphoglucomutase, and uridine diphosphoglucose pyrophosphorylase. The oxygen-18 at the alpha beta-bridge position of the labeled UDP-Glc has been shown to cause a 0.014 ppm upfield chemical shift in the 31P NMR spectrum of both the alpha- and beta-phosphorus atoms in UDP-Glc relative to the unlabeled compound. The chemical shift induced by each of the beta-nonbridge oxygen-18 atoms was 0.030 ppm. Incubation of (beta-/sup 18/O/sub 2/, alpha beta-/sup 18/O)UDP-Glc with sucrose synthetase in the presence and absence of 2,5-anhydromannitol did not result in any significant exchange of an oxygen-18 from the beta-nonbridge position to the anomeric oxygen of the glucose moiety. It can thus be concluded that either sucrose synthetase does not catalyze the cleavage of the scissile carbon-oxygen bond of UDP-Glc in the absence of fructose or, alternatively, the beta-phosphoryl group of the newly formed UDP is rotationally immobilized.

  11. Dual targeting is the rule for organellar aminoacyl-tRNA synthetases in Arabidopsis thaliana

    PubMed Central

    Duchêne, Anne-Marie; Giritch, Anatoli; Hoffmann, Beate; Cognat, Valérie; Lancelin, Dominique; Peeters, Nemo M.; Zaepfel, Marlyse; Maréchal-Drouard, Laurence; Small, Ian D.

    2005-01-01

    In plants, protein synthesis occurs in the cytosol, mitochondria, and plastids. Each compartment requires a full set of tRNAs and aminoacyl-tRNA synthetases. We have undertaken a systematic analysis of the targeting of organellar aminoacyl-tRNA synthetases in the model plant Arabidopsis thaliana. Dual targeting appeared to be a general rule. Among the 24 identified organellar aminoacyl-tRNA synthetases (aaRSs), 15 (and probably 17) are shared between mitochondria and plastids, and 5 are shared between cytosol and mitochondria (one of these aaRSs being present also in chloroplasts). Only two were shown to be uniquely chloroplastic and none to be uniquely mitochondrial. Moreover, there are no examples where the three aaRS genes originating from the three ancestral genomes still coexist. These results indicate that extensive exchange of aaRSs has occurred during evolution and that many are now shared between two or even three compartments. The findings have important implications for studies of the translation machinery in plants and on protein targeting and gene transfer in general. PMID:16251277

  12. 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. PMID:27234879

  13. Structural basis for the binding of succinate to succinyl-CoA synthetase.

    PubMed

    Huang, Ji; Fraser, Marie E

    2016-08-01

    Succinyl-CoA synthetase catalyzes the only step in the citric acid cycle that provides substrate-level phosphorylation. Although the binding sites for the substrates CoA, phosphate, and the nucleotides ADP and ATP or GDP and GTP have been identified, the binding site for succinate has not. To determine this binding site, pig GTP-specific succinyl-CoA synthetase was crystallized in the presence of succinate, magnesium ions and CoA, and the structure of the complex was determined by X-ray crystallography to 2.2 Å resolution. Succinate binds in the carboxy-terminal domain of the β-subunit. The succinate-binding site is near both the active-site histidine residue that is phosphorylated in the reaction and the free thiol of CoA. The carboxy-terminal domain rearranges when succinate binds, burying this active site. However, succinate is not in position for transfer of the phosphoryl group from phosphohistidine. Here, it is proposed that when the active-site histidine residue has been phosphorylated by GTP, the phosphohistidine displaces phosphate and triggers the movement of the carboxylate of succinate into position to be phosphorylated. The structure shows why succinyl-CoA synthetase is specific for succinate and does not react appreciably with citrate nor with the other C4-dicarboxylic acids of the citric acid cycle, fumarate and oxaloacetate, but shows some activity with L-malate. PMID:27487822

  14. Orthogonal Protein Assembly on DNA Nanostructures Using Relaxases.

    PubMed

    Sagredo, Sandra; Pirzer, Tobias; Aghebat Rafat, Ali; Goetzfried, Marisa A; Moncalian, Gabriel; Simmel, Friedrich C; de la Cruz, Fernando

    2016-03-18

    DNA-binding proteins are promising reagents for the sequence-specific modification of DNA-based nanostructures. Here, we investigate the utility of a series of relaxase proteins-TrwC, TraI, and MobA-for nanofunctionalization. Relaxases are involved in the conjugative transfer of plasmids between bacteria, and bind to their DNA target sites via a covalent phosphotyrosine linkage. We study the binding of the relaxases to two standard DNA origami structures-rodlike six-helix bundles and flat rectangular origami sheets. We find highly orthogonal binding of the proteins with binding yields of 40-50 % per binding site, which is comparable to other functionalization methods. The yields differ for the two origami structures and also depend on the position of the binding sites. Due to their specificity for a single-stranded DNA target, their orthogonality, and their binding properties, relaxases are a uniquely useful addition to the toolbox available for the modification of DNA nanostructures with proteins. PMID:26915475

  15. Xylonucleic acid: synthesis, structure, and orthogonal pairing properties

    PubMed Central

    Maiti, Mohitosh; Maiti, Munmun; Knies, Christine; Dumbre, Shrinivas; Lescrinier, Eveline; Rosemeyer, Helmut; Ceulemans, Arnout; Herdewijn, Piet

    2015-01-01

    There is a common interest for studying xeno-nucleic acid systems in the fields of synthetic biology and the origin of life, in particular, those with an engineered backbone and possessing novel properties. Along this line, we have investigated xylonucleic acid (XyloNA) containing a potentially prebiotic xylose sugar (a 3′-epimer of ribose) in its backbone. Herein, we report for the first time the synthesis of four XyloNA nucleotide building blocks and the assembly of XyloNA oligonucleotides containing all the natural nucleobases. A detailed investigation of pairing and structural properties of XyloNAs in comparison to DNA/RNA has been performed by thermal UV-melting, CD, and solution state NMR spectroscopic studies. XyloNA has been shown to be an orthogonal self-pairing system which adopts a slightly right-handed extended helical geometry. Our study on one hand, provides understanding for superior structure-function (-pairing) properties of DNA/RNA over XyloNA for selection as an informational polymer in the prebiotic context, while on the other hand, finds potential of XyloNA as an orthogonal genetic system for application in synthetic biology. PMID:26175047

  16. 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.

  17. Orthogonal Cherenkov sound in spin-orbit coupled systems

    NASA Astrophysics Data System (ADS)

    Smirnov, Sergey

    2015-06-01

    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.

  18. 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.

  19. 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.

  20. 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

  1. 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

  2. Improved analytical model for residual stress prediction in orthogonal cutting

    NASA Astrophysics Data System (ADS)

    Qi, Zhaoxu; Li, Bin; Xiong, Liangshan

    2014-09-01

    The analytical model of residual stress in orthogonal cutting proposed by Jiann is an important tool for residual stress prediction in orthogonal cutting. In application of the model, a problem of low precision of the surface residual stress prediction is found. By theoretical analysis, several shortages of Jiann's model are picked out, including: inappropriate boundary conditions, unreasonable calculation method of thermal stress, ignorance of stress constraint and cyclic loading algorithm. These shortages may directly lead to the low precision of the surface residual stress prediction. To eliminate these shortages and make the prediction more accurate, an improved model is proposed. In this model, a new contact boundary condition between tool and workpiece is used to make it in accord with the real cutting process; an improved calculation method of thermal stress is adopted; a stress constraint is added according to the volumeconstancy of plastic deformation; and the accumulative effect of the stresses during cyclic loading is considered. At last, an experiment for measuring residual stress in cutting AISI 1045 steel is conducted. Also, Jiann's model and the improved model are simulated under the same conditions with cutting experiment. The comparisons show that the surface residual stresses predicted by the improved model is closer to the experimental results than the results predicted by Jiann's model.

  3. Improved analytical model for residual stress prediction in orthogonal cutting

    NASA Astrophysics Data System (ADS)

    Qi, Zhaoxu; Li, Bin; Xiong, Liangshan

    2014-09-01

    The analytical model of residual stress in orthogonal cutting proposed by Jiann is an important tool for residual stress prediction in orthogonal cutting. In application of the model, a problem of low precision of the surface residual stress prediction is found. By theoretical analysis, several shortages of Jiann's model are picked out, including: inappropriate boundary conditions, unreasonable calculation method of thermal stress, ignorance of stress constraint and cyclic loading algorithm. These shortages may directly lead to the low precision of the surface residual stress prediction. To eliminate these shortages and make the prediction more accurate, an improved model is proposed. In this model, a new contact boundary condition between tool and workpiece is used to make it in accord with the real cutting process; an improved calculation method of thermal stress is adopted; a stress constraint is added according to the volume-constancy of plastic deformation; and the accumulative effect of the stresses during cyclic loading is considered. At last, an experiment for measuring residual stress in cutting AISI 1045 steel is conducted. Also, Jiann's model and the improved model are simulated under the same conditions with cutting experiment. The comparisons show that the surface residual stresses predicted by the improved model is closer to the experimental results than the results predicted by Jiann's model.

  4. Orthogonal Connectivity Factorization: Interpretable Decomposition of Variability in Correlation Matrices.

    PubMed

    Hyvärinen, Aapo; Hirayama, Jun-ichiro; Kiviniemi, Vesa; Kawanabe, Motoaki

    2016-03-01

    In many multivariate time series, the correlation structure is nonstationary, that is, it changes over time. The correlation structure may also change as a function of other cofactors, for example, the identity of the subject in biomedical data. A fundamental approach for the analysis of such data is to estimate the correlation structure (connectivities) separately in short time windows or for different subjects and use existing machine learning methods, such as principal component analysis (PCA), to summarize or visualize the changes in connectivity. However, the visualization of such a straightforward PCA is problematic because the ensuing connectivity patterns are much more complex objects than, say, spatial patterns. Here, we develop a new framework for analyzing variability in connectivities using the PCA approach as the starting point. First, we show how to analyze and visualize the principal components of connectivity matrices by a tailor-made rank-two matrix approximation in which we use the outer product of two orthogonal vectors. This leads to a new kind of transformation of eigenvectors that is particularly suited for this purpose and often enables interpretation of the principal component as connectivity between two groups of variables. Second, we show how to incorporate the orthogonality and the rank-two constraint in the estimation of PCA itself to improve the results. We further provide an interpretation of these methods in terms of estimation of a probabilistic generative model related to blind separation of dependent sources. Experiments on brain imaging data give very promising results. PMID:26735746

  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. Empirical Orthogonal Functions of the Australian Regional Ionosphere

    NASA Astrophysics Data System (ADS)

    Bouya, Z.; Terkildsen, M.; Francis, M.

    2011-12-01

    Understanding the natural background variability of the ionosphere is required for investigations of the ionospheric response to external perturbations. Decomposition of ionospheric observations into a set of spatiotemporal structures allows the separation of the anomalous response of the ionosphere from the background variability. This paper presents preliminary results of the analysis of Australian regional Total Electron Content (TEC) using an Empirical Orthogonal Function (EOF) technique to distinguish the variability of the ionosphere caused by the solar and geomagnetic activity. TEC data at evenly distributed grid points was estimated using the Spherical Cap Harmonic Analysis (SCHA) method, applied to GPS data collected from the Australian Regional GPS Network (ARGN). This approach yields more detailed maps of regional TEC variation than can be obtained using global models. EOF analysis was used to decompose the TEC data set into a series of orthogonal Eigenfunction (EOF base function) and associated coefficients. The base function represents the variation in TEC with latitude and longitude. The coefficients represent the variation with time. Analysis of a year worth of data from 2010 yielded the following results: The first 5 EOFs describe 95% of the total TEC variance. The equatorial zone dominates in the spatial structure of the first EOF. This mode has a distinct diurnal and seasonal evolution its proportion is ~65% and highly correlates with the solar activity. Keywords: EOF, Equatorial Anomaly, Regional, SCHA, TEC, GPS, Australia .

  7. Control of the Synthesis of Fatty-Acid Synthetase in Rat Liver by Insulin, Glucagon, and Adenosine 3′:5′ Cyclic Monophosphate

    PubMed Central

    Lakshmanan, M. R.; Nepokroeff, Carl M.; Porter, John W.

    1972-01-01

    The usual increase in the activity of liver fatty-acid synthetase that occurs on refeeding of a fat-free diet to previously fasted rats is abolished in diabetic animals. Insulin specifically restores this increase by enhancement of the rate of synthesis of fatty-acid synthetase. However, glucagon and cyclic AMP inhibit the increase in the activity of fatty-acid synthetase. Therefore, the concentration of fatty-acid synthetase in rat liver is under the control of the relative concentrations of insulin and glucagon. PMID:4345502

  8. Isolation and Partial Characterization of Temperature-Sensitive Escherichia coli Mutants with Altered Leucyl- and Seryl-Transfer Ribonucleic Acid Synthetases

    PubMed Central

    Low, B.; Gates, F.; Goldstein, T.; Söll, D.

    1971-01-01

    Two temperature-sensitive mutants of Escherichia coli have been found in which the conditional growth is a result of a thermosensitive leucyl-transfer ribonucleic acid (tRNA) synthetase and seryl-tRNA synthetase, respectively. The corresponding genetic loci, leuS and serS, cotransduce with lip and serC, respectively. As a result of the mutationally altered leucyl-tRNA synthetase, some leucine-, valine-, and isoleucine-forming enzymes were derepressed. Thus, leucyl-tRNA synthetase is involved in the repression of the enzymes needed for the synthesis of branched-chain amino acids. PMID:4942762

  9. Activation of 2',5'-oligoadenylate synthetase activity on induction of HL-60 leukemia cell differentiation.

    PubMed Central

    Schwartz, E L; Nilson, L A

    1989-01-01

    A 27-fold increase in 2',5'-oligoadenylate synthetase activity, an enzyme associated with the antiproliferative actions of interferon (IFN), was observed after treatment of HL-60 human leukemia cells with dimethyl sulfoxide (DMSO), an inducer of granulocytic differentiation of the cells. Enzyme activity was elevated after 24 h of exposure to DMSO, was maximal at 48 hours, and declined thereafter. A comparable increase was observed after treatment with 1 U of alpha interferon (IFN-alpha) per ml or 8 U of beta interferon (IFN-beta) per ml. Elevated levels of expression of other IFN-inducible genes, including type I histocompatibility antigen (HLA-B) mRNA and 2',5'-oligoadenylate phosphodiesterase activity, were also observed with DMSO treatment. DMSO-treated HL-60 cells had an increased amount of a 1.8-kilobase mRNA for oligoadenylate [oligo(A)] synthetase when compared with that of control cells; both DMSO- and IFN-treated HL-60 cells also expressed 1.6-, 3.4-, and 4.3-kilobase mRNA. The increase in both oligo(A) synthetase activity and mRNA levels was inhibited by polyclonal antiserum to human IFN-alpha; however, no IFN-alpha mRNA could be detected in the cells. Antiserum to IFN-beta or gamma interferon (IFN-gamma) had no effect on oligo(A) synthetase expression or activity nor was there any detectable IFN-beta 1 or IFN-beta 2 mRNA in the cells. The anti-IFN-alpha serum did not block the elevation of HLA-B mRNA in DMSO-treated cells. These observations suggest that the increased expression of oligo(A) synthetase in DMSO-treated cells may be mediated by the release of an IFN-alpha-like factor; however, the levels of any IFN-alpha mRNA produced in the cells were extremely low. Images PMID:2476665

  10. Biosynthesis of Polymyxins B, E, and P Using Genetically Engineered Polymyxin Synthetases in the Surrogate Host Bacillus subtilis.

    PubMed

    Kim, Se-Yu; Park, Soo-Young; Choi, Soo-Keun; Park, Seung-Hwan

    2015-07-01

    The development of diverse polymyxin derivatives is needed to solve the toxicity and resistance problems of polymyxins. However, no platform has generated polymyxin derivatives by genetically engineering a polymyxin synthetase, which is a nonribosomal peptide synthetase. In this study, we present a two-step approach for the construction of engineered polymyxin synthetases by substituting the adenylation (A) domains of polymyxin A synthetase, which is encoded by the pmxABCDE gene cluster of Paenibacillus polymyxa E681. First, the seventh L-threonine-specific A-domain region in pmxA was substituted with the Lleucine- specific A-domain region obtained from P. polymyxa ATCC21830 to make polymyxin E synthetase, and then the sixth D-leucine-specific A-domain region (A6-D-Leu-domain) was substituted with the D-phenylalanine-specific A-domain region (A6-D-Phe-domain) obtained from P. polymyxa F4 to make polymyxin B synthetase. This step was performed in Escherichia coli on a pmxA-containing fosmid, using the lambda Red recombination system and the sacB gene as a counter-selectable marker. Next, the modified pmxA gene was fused to pmxBCDE on the chromosome of Bacillus subtilis BSK4dA, and the resulting recombinant strains BSK4-PB and BSK4-PE were confirmed to produce polymyxins B and E, respectively. We also succeeded in constructing the B. subtilis BSK4-PP strain, which produces polymyxin P, by singly substituting the A6-D-Leu-domain with the A6-D-Phe-domain. This is the first report in which polymyxin derivatives were generated by genetically engineering polymyxin synthetases. The two recombinant B. subtilis strains will be useful for improving the commercial production of polymyxins B and E, and they will facilitate the generation of novel polymyxin derivatives. PMID:26059516

  11. Root of the universal tree of life based on ancient aminoacyl-tRNA synthetase gene duplications.

    PubMed

    Brown, J R; Doolittle, W F

    1995-03-28

    Universal trees based on sequences of single gene homologs cannot be rooted. Iwabe et al. [Iwabe, N., Kuma, K.-I., Hasegawa, M., Osawa, S. & Miyata, T. (1989) Proc. Natl. Acad. Sci. USA 86, 9355-9359] circumvented this problem by using ancient gene duplications that predated the last common ancestor of all living things. Their separate, reciprocally rooted gene trees for elongation factors and ATPase subunits showed Bacteria (eubacteria) as branching first from the universal tree with Archaea (archaebacteria) and Eucarya (eukaryotes) as sister groups. Given its topical importance to evolutionary biology and concerns about the appropriateness of the ATPase data set, an evaluation of the universal tree root using other ancient gene duplications is essential. In this study, we derive a rooting for the universal tree using aminoacyl-tRNA synthetase genes, an extensive multigene family whose divergence likely preceded that of prokaryotes and eukaryotes. An approximately 1600-bp conserved region was sequenced from the isoleucyl-tRNA synthetases of several species representing deep evolutionary branches of eukaryotes (Nosema locustae), Bacteria (Aquifex pyrophilus and Thermotoga maritima) and Archaea (Pyrococcus furiosus and Sulfolobus acidocaldarius). In addition, a new valyl-tRNA synthetase was characterized from the protist Trichomonas vaginalis. Different phylogenetic methods were used to generate trees of isoleucyl-tRNA synthetases rooted by valyl- and leucyl-tRNA synthetases. All isoleucyl-tRNA synthetase trees showed Archaea and Eucarya as sister groups, providing strong confirmation for the universal tree rooting reported by Iwabe et al. As well, there was strong support for the monophyly (sensu Hennig) of Archaea. The valyl-tRNA synthetase gene from Tr. vaginalis clustered with other eukaryotic ValRS genes, which may have been transferred from the mitochondrial genome to the nuclear genome, suggesting that this amitochondrial trichomonad once harbored an

  12. Coherent vorticity extraction in resistive drift-wave turbulence: Comparison of orthogonal wavelets versus proper orthogonal decomposition

    SciTech Connect

    Futatani, S.; Bos, W.J.T.; Del-Castillo-Negrete, Diego B; Schneider, Kai; Benkadda, S.; Farge, Marie

    2011-01-01

    We assess two techniques for extracting coherent vortices out of turbulent flows: the wavelet based Coherent Vorticity Extraction (CVE) and the Proper Orthogonal Decomposition (POD). The former decomposes the flow field into an orthogonal wavelet representation and subsequent thresholding of the coefficients allows one to split the flow into organized coherent vortices with non-Gaussian statistics and an incoherent random part which is structureless. POD is based on the singular value decomposition and decomposes the flow into basis functions which are optimal with respect to the retained energy for the ensemble average. Both techniques are applied to direct numerical simulation data of two-dimensional drift-wave turbulence governed by Hasegawa Wakatani equation, considering two limit cases: the quasi-hydrodynamic and the quasi-adiabatic regimes. The results are compared in terms of compression rate, retained energy, retained enstrophy and retained radial flux, together with the enstrophy spectrum and higher order statistics. (c) 2010 Published by Elsevier Masson SAS on behalf of Academie des sciences.

  13. Three Dimensional Wind Speed and Flux Measurement over a Rain-fed Soybean Field Using Orthogonal and Non-orthogonal Sonic Anemometer Designs

    NASA Astrophysics Data System (ADS)

    Thomas, T.; Suyker, A.; Burba, G. G.; Billesbach, D.

    2014-12-01

    The eddy covariance method for estimating fluxes of trace gases, energy and momentum in the constant flux layer above a plant canopy fundamentally relies on accurate measurements of the vertical wind speed. This wind speed is typically measured using a three dimensional ultrasonic anemometer. These anemometers incorporate designs with transducer sets that are aligned either orthogonally or non-orthogonally. Previous studies comparing the two designs suggest differences in measured 3D wind speed components, in particular vertical wind speed, from the non-orthogonal transducer relative to the orthogonal design. These differences, attributed to additional flow distortion caused by the non-orthogonal transducer arrangement, directly affect fluxes of trace gases, energy and momentum. A field experiment is being conducted over a rain-fed soybean field at the AmeriFlux site (US-Ne3) near Mead, Nebraska. In this study, ultrasonic anemometers featuring orthogonal transducer sets (ATI Vx Probe) and non-orthogonal transducer sets (Gill R3-100) collect high frequency wind vector and sonic temperature data. Sensible heat and momentum fluxes and other key sonic performance data are evaluated based on environmental parameters including wind speed, wind direction, temperature, and angle of attack. Preliminary field experiment results are presented.

  14. Application of neural networks with orthogonal activation functions in control of dynamical systems

    NASA Astrophysics Data System (ADS)

    Nikolić, Saša S.; Antić, Dragan S.; Milojković, Marko T.; Milovanović, Miroslav B.; Perić, Staniša Lj.; Mitić, Darko B.

    2016-04-01

    In this article, we present a new method for the synthesis of almost and quasi-orthogonal polynomials of arbitrary order. Filters designed on the bases of these functions are generators of generalised quasi-orthogonal signals for which we derived and presented necessary mathematical background. Based on theoretical results, we designed and practically implemented generalised first-order (k = 1) quasi-orthogonal filter and proved its quasi-orthogonality via performed experiments. Designed filters can be applied in many scientific areas. In this article, generated functions were successfully implemented in Nonlinear Auto Regressive eXogenous (NARX) neural network as activation functions. One practical application of the designed orthogonal neural network is demonstrated through the example of control of the complex technical non-linear system - laboratory magnetic levitation system. Obtained results were compared with neural networks with standard activation functions and orthogonal functions of trigonometric shape. The proposed network demonstrated superiority over existing solutions in the sense of system performances.

  15. One method for calculating flux-MMF relationship of orthogonal-core

    SciTech Connect

    Tajima, Katsubumi; Kaga, Akio; Anazawa, Yoshihisa . Dept. of Electrical and Electronics Engineering); Ichinokura, Osamu . Dept. of Electrical Engineering)

    1993-11-01

    The orthogonal-core has various applications, e.g. as a variable inductor, a parametric transformer and a DC-AC converter. This paper describes one method for calculating the flux-MMF relationship of an orthogonal-core. The calculation is based on a 3-dimensional magnetic circuit model of the orthogonal-core. The model is derived by dividing the orthogonal-core, inclusive of the surrounding region, into elements comprising a 3-dimensional magnetic circuit. Using this model, the authors can compute the flux-MMF relationship of the orthogonal core with arbitrary dimensions from the B-H characteristic of the core material. The calculation method presented here is useful for optimum design of devices using an orthogonal-core.

  16. Ensemble forecast of typhoon generated by orthogonal conditional nonlinear optimal perturbations

    NASA Astrophysics Data System (ADS)

    Huo, Zhenhua; Duan, Wansuo; Zhou, Feifan

    2016-04-01

    Orthogonal conditional nonlinear optimal perturbations (CNOPs) are the initial perturbations that have the largest impact on the forecast results in orthogonal subspaces of the initial perturbation space. Previous studies demonstrate the successful application of orthogonal CNOPs in ensemble forecasting. And further analysis indicates that orthogonal CNOPs may be more adapt to the prediction of strong events, among which typhoon events occur in the tropical or subtropical areas where the diabatic physical processes is very important and has strong nonlinear behavior. For these reasons, this paper focuses on the application of orthogonal CNOPs in ensemble forecast of typhoon. In this study, orthogonal CNOPs, orthogonal singular vectors (SVs), bred vectors (BVs) and random perturbations (RPs) are applied for typhoon ensemble forecasts using MM5 model. The results show that, for typhoons Matsa in 2005 and Sepat in 2007, ensemble forecasts generated by orthogonal CNOPs greatly improve the control forecast, successfully predicts the landing location of Matsa, and gives the warning information of the landing of Sepat. In detail, for the ensemble mean associated with orthogonal CNOPs, the averaging track forecast error over 5 days is decreased by 45.58 km for Matsa and 87.8 km for Sepat, compared with control forecast. However, ensemble forecasts generated by other three methods could not successfully predict the landing location of Matsa and give the warning information of the landing of Sepat. Compared with orthogonal SVs, BVs and RPs, ensemble forecasts generated by orthogonal CNOPs corresponds to the largest ensemble spread, improves the control forecast at the largest extent, and best samples the distribution of initial analysis errors. All these results show that orthogonal CNOPs may provide another useful technique for ensemble forecast of typhoon.

  17. State diagram of an orthogonal spin transfer spin valve device

    SciTech Connect

    Ye, Li; Wolf, Georg; Pinna, Daniele; Chaves-O'Flynn, Gabriel D.; Kent, Andrew D.

    2015-05-21

    We present the switching characteristics of a spin-transfer device that incorporates a perpendicularly magnetized spin-polarizing layer with an in-plane magnetized free and fixed magnetic layer, known as an orthogonal spin transfer spin valve device. This device shows clear switching between parallel (P) and antiparallel (AP) resistance states and the reverse transition (AP → P) for both current polarities. Further, hysteretic transitions are shown to occur into a state with a resistance intermediate between that of the P and AP states, again for both current polarities. These unusual spin-transfer switching characteristics can be explained within a simple macrospin model that incorporates thermal fluctuations and considers a spin-polarized current that is tilted with respect to the free layer's plane, due to the presence of the spin-transfer torque from the polarizing layer.

  18. Orthogonally referenced integrated ensemble for navigation and timing

    SciTech Connect

    Smith, Stephen Fulton; Moore, James Anthony

    2014-04-01

    An orthogonally referenced integrated ensemble for navigation and timing includes a dual-polyhedral oscillator array, including an outer sensing array of oscillators and an inner clock array of oscillators situated inside the outer sensing array. The outer sensing array includes a first pair of sensing oscillators situated along a first axis of the outer sensing array, a second pair of sensing oscillators situated along a second axis of the outer sensing array, and a third pair of sensing oscillators situated along a third axis of the outer sensing array. The inner clock array of oscillators includes a first pair of clock oscillators situated along a first axis of the inner clock array, a second pair of clock oscillators situated along a second axis of the inner clock array, and a third pair of clock oscillators situated along a third axis of the inner clock array.

  19. Orthogonally referenced integrated ensemble for navigation and timing

    SciTech Connect

    Smith, Stephen Fulton; Moore, James Anthony

    2013-02-26

    An orthogonally referenced integrated ensemble for navigation and timing includes a dual-polyhedral oscillator array, including an outer sensing array of oscillators and an inner clock array of oscillators situated inside the outer sensing array. The outer sensing array includes a first pair of sensing oscillators situated along a first axis of the outer sensing array, a second pair of sensing oscillators situated along a second axis of the outer sensing array, and a third pair of sensing oscillators situated along a third axis of the outer sensing array. The inner clock array of oscillators includes a first pair of clock oscillators situated along a first axis of the inner clock array, a second pair of clock oscillators situated along a second axis of the inner clock array, and a third pair of clock oscillators situated along a third axis of the inner clock array.

  20. Extraction optimization of carbohydrate compound from Huangqi using orthogonal design.

    PubMed

    Zhou, Shen-kang; Bi, Tie-Nan; Xu, Yun-Feng; Zhang, Rui-Li; Yang, Mei-Juan

    2013-07-01

    The effect of extraction time, extraction temperature and time on the yield of Huangqi carbohydrate compound (HQCC) was investigated using single factor and orthogonal experiment design. The influence by the parameters on the extraction yields of carbohydrate compound decreased in the order of: C (extraction number)>A (extraction time)>B (extraction temperature) according to the R values. Based on this analysis, and considering the carbohydrate compound extraction efficiency, the cost of energy and the feasibility of experiment, the optimum conditions of extraction were therefore determined as follows: extraction time 120min, extraction temperature 80°C, and extraction number 4. Oral administration of HQCC reduced lipid peroxidation level and enhanced antioxidant enzymes activities in gastric mucosa. In addition, HQCC reduced the serum IL-8 and TNF-α levels. In conclusion, these data reveal that HQCC promotes regeneration of damaged gastric mucosa, probably through its antioxidative mechanism. PMID:23541555

  1. Vectorial strain gauge method using single flexible orthogonal polydimethylsiloxane gratings

    PubMed Central

    Guo, Hao; Tang, Jun; Qian, Kun; Tsoukalas, Dimitris; Zhao, Miaomiao; Yang, Jiangtao; Zhang, Binzhen; Chou, Xiujian; Liu, Jun; Xue, Chenyang; Zhang, Wendong

    2016-01-01

    A vectorial strain gauge method using a single sensing element is reported based on the double-sided polydimethylsiloxane (PDMS) Fraunhofer diffraction gratings structures. Using O2 plasma treatment steps, orthogonal wrinkled gratings were fabricated on both sides of a pre-strained PDMS film. Diffracted laser spots from this structure have been used to experimentally demonstrate, that any applied strain can be quantitatively characterized in both the x and y directions with an error of less than 0.6% and with a gauge factor of approximately 10. This simple and low cost technology which is completely different from the traditional vectorial strain gauge method, can be applied to surface vectorial strain measurement and multi-axis integrated mechanical sensors. PMID:27005493

  2. Improved orthogonal frequency division multiplexing communications through advanced coding

    NASA Astrophysics Data System (ADS)

    Westra, Jeffrey; Patti, John

    2005-08-01

    Orthogonal Frequency Division Multiplexing (OFDM) is a communications technique that transmits a signal over multiple, evenly spaced, discrete frequency bands. OFDM offers some advantages over traditional, single-carrier modulation techniques, such as increased immunity to inter-symbol interference. For this reason OFDM is an attractive candidate for sensor network application; it has already been included in several standards, including Digital Audio Broadcast (DAB); digital television standards in Europe, Japan and Australia; asymmetric digital subscriber line (ASDL); and wireless local area networks (WLAN), specifically IEEE 802.11a. Many of these applications currently make use of a standard convolutional code with Viterbi decoding to perform forward error correction (FEC). Replacing such convolutional codes with advanced coding techniques using iterative decoding, such as Turbo codes, can substantially improve the performance of the OFDM communications link. This paper demonstrates such improvements using the 802.11a wireless LAN standard.

  3. Vectorial strain gauge method using single flexible orthogonal polydimethylsiloxane gratings.

    PubMed

    Guo, Hao; Tang, Jun; Qian, Kun; Tsoukalas, Dimitris; Zhao, Miaomiao; Yang, Jiangtao; Zhang, Binzhen; Chou, Xiujian; Liu, Jun; Xue, Chenyang; Zhang, Wendong

    2016-01-01

    A vectorial strain gauge method using a single sensing element is reported based on the double-sided polydimethylsiloxane (PDMS) Fraunhofer diffraction gratings structures. Using O2 plasma treatment steps, orthogonal wrinkled gratings were fabricated on both sides of a pre-strained PDMS film. Diffracted laser spots from this structure have been used to experimentally demonstrate, that any applied strain can be quantitatively characterized in both the x and y directions with an error of less than 0.6% and with a gauge factor of approximately 10. This simple and low cost technology which is completely different from the traditional vectorial strain gauge method, can be applied to surface vectorial strain measurement and multi-axis integrated mechanical sensors. PMID:27005493

  4. Integrated power divider/combiner at hybrid orthogonal junctions

    NASA Astrophysics Data System (ADS)

    El Sherif, Mohamed H.; Ahmed, Osman S.; Bakr, Mohamed H.

    2014-03-01

    We propose an integrated power divider/combiner at the interface between silicon nanowire and plasmonic slot waveguide (PSW). The proposed configuration facilitates light access and manipulation in planar nano-plasmonic circuits. The light is incident from a standard silicon nanowire to be accessed by a nano-plasmonic circuit providing subwavelength confinement. The structure overcomes the losses associated with long distance light propagation in nanoplasmonic splitters as coupling and splitting are performed at the same interface with minimal losses. Two PSWs placed orthogonally to the silicon nanowire forming hybrid junctions are exploited for the power dividing/combining functionality. The power splitter has been analyzed using the finite difference time domain (FDTD) numerical method. The ultra-compact proposed device provides wide-band power splitting functionality. A splitting of 34.7%, over most of the wavelength spectrum from 0.8 μm - 2.5 μm, is demonstrated.

  5. Rational design of orthogonal libraries of protein coding genes.

    PubMed

    Ryan, Daniel; Papamichail, Dimitris

    2013-05-17

    Array-based oligonucleotide synthesis technologies provide access to thousands of custom-designed sequence variants at low cost. Large-scale synthesis and high-throughput assays have become valuable experimental tools to study in detail the interplay between sequence and function. We have developed a methodology and corresponding algorithms for the design of diverse protein coding gene libraries, to exploit the potential of multiplex synthesis and help elucidate the effects of codon utilization and other factors in gene expression. Using our algorithm, we have computationally designed gene libraries with hundreds to thousands of orthogonal codon usage variants, uniformly exploring the design space of codon utilization, while demanding only a small fraction of the synthesis cost that would be required if these variants were synthesized independently. PMID:23654273

  6. Sequence-defined polymers via orthogonal allyl acrylamide building blocks.

    PubMed

    Porel, Mintu; Alabi, Christopher A

    2014-09-24

    Biological systems have long recognized the importance of macromolecular diversity and have evolved efficient processes for the rapid synthesis of sequence-defined biopolymers. However, achieving sequence control via synthetic methods has proven to be a difficult challenge. Herein we describe efforts to circumvent this difficulty via the use of orthogonal allyl acrylamide building blocks and a liquid-phase fluorous support for the de novo design and synthesis of sequence-specific polymers. We demonstrate proof-of-concept via synthesis and characterization of two sequence-isomeric 10-mer polymers. (1)H NMR and LCMS were used to confirm their chemical structure while tandem MS was used to confirm sequence identity. Further validation of this methodology was provided via the successful synthesis of a sequence-specific 16-mer polymer incorporating nine different monomers. This strategy thus shows promise as an efficient approach for the assembly of sequence-specific functional polymers. PMID:25204618

  7. Asymptotic formulae for the zeros of orthogonal polynomials

    SciTech Connect

    Badkov, V M

    2012-09-30

    Let p{sub n}(t) be an algebraic polynomial that is orthonormal with weight p(t) on the interval [-1, 1]. When p(t) is a perturbation (in certain limits) of the Chebyshev weight of the first kind, the zeros of the polynomial p{sub n}( cos {tau}) and the differences between pairs of (not necessarily consecutive) zeros are shown to satisfy asymptotic formulae as n{yields}{infinity}, which hold uniformly with respect to the indices of the zeros. Similar results are also obtained for perturbations of the Chebyshev weight of the second kind. First, some preliminary results on the asymptotic behaviour of the difference between two zeros of an orthogonal trigonometric polynomial, which are needed, are established. Bibliography: 15 titles.

  8. Antioxidant peptides from corn gluten meal: Orthogonal design evaluation.

    PubMed

    Zhou, Cunshan; Hu, Jiali; Ma, Haile; Yagoub, Abu ElGasim A; Yu, Xiaojie; Owusu, John; Ma, Haiyan; Qin, Xiaopei

    2015-11-15

    Protamex catalyzed corn gluten meal (CGM) hydrolysis peptides (CHP) were prepared. Orthogonal design L16 (4(5)) was used to optimize processing variables of CGM concentration, CGM heat pretreatment (121 °C) time, and enzymolysis pH, temperature, and time. Degree of hydrolysis (DH), undigested residue ratio, molecular weight (MW) distribution and DPPH radical inhibition were selected as analysis indicators. Optimum variables were CGM concentration of 18%, heat pretreatment time of 40 min, and enzymolysis pH, temperature and time of 7.5, 55 °C and 24h, respectively. Verification test showed that CHP IC50 for scavenging hydroxyl radical was the best and then followed by reducing power. Oligopeptides improved after hydrolysis at the expense of di- and tripeptides, suggesting formation of soluble aggregates from low MW peptides. The increase in the DH, oligopeptides, Alanyl-Tyrosine, and antioxidant free amino acids coincided with the improvement in the antioxidant activity of CHP. PMID:25977026

  9. Vectorial strain gauge method using single flexible orthogonal polydimethylsiloxane gratings

    NASA Astrophysics Data System (ADS)

    Guo, Hao; Tang, Jun; Qian, Kun; Tsoukalas, Dimitris; Zhao, Miaomiao; Yang, Jiangtao; Zhang, Binzhen; Chou, Xiujian; Liu, Jun; Xue, Chenyang; Zhang, Wendong

    2016-03-01

    A vectorial strain gauge method using a single sensing element is reported based on the double-sided polydimethylsiloxane (PDMS) Fraunhofer diffraction gratings structures. Using O2 plasma treatment steps, orthogonal wrinkled gratings were fabricated on both sides of a pre-strained PDMS film. Diffracted laser spots from this structure have been used to experimentally demonstrate, that any applied strain can be quantitatively characterized in both the x and y directions with an error of less than 0.6% and with a gauge factor of approximately 10. This simple and low cost technology which is completely different from the traditional vectorial strain gauge method, can be applied to surface vectorial strain measurement and multi-axis integrated mechanical sensors.

  10. The Full Classification of Orthogonal Easy Quantum Groups

    NASA Astrophysics Data System (ADS)

    Raum, Sven; Weber, Moritz

    2016-02-01

    We study easy quantum groups, a combinatorial class of orthogonal quantum groups introduced by Banica-Speicher in 2009. We show that there is a countable descending chain of easy quantum groups interpolating between Bichon's free wreath product with the permutation group S n and a semi-direct product of a permutation action of S n on a free product. This reveals a series of new commutation relations interpolating between a free product construction and the tensor product. Furthermore, we prove a dichotomy result saying that every hyperoctahedral easy quantum group is either part of our new interpolating series of quantum groups or belongs to a class of semi-direct product quantum groups recently studied by the authors. This completes the classification of easy quantum groups. We also study combinatorial and operator algebraic aspects of the new interpolating series.

  11. Enrichment and characterization of the mRNAs of four aminoacyl-tRNA synthetases from yeast.

    PubMed Central

    Sellami, M; Rether, B; Gangloff, J; Ebel, J P; Bonnet, J

    1983-01-01

    We have partially purified the messenger RNAs for yeast arginyl-, aspartyl-, valyl-, alpha and beta subunits of phenylalanyl-tRNA synthetases in order to study their biosynthesis and ultimately, to isolate their genes. Sucrose gradient fractionation of poly U-Sepharose selected mRNAs resulted in a ten fold enrichment of the in vitro translation activity of these mRNAs. The translation products of messenger RNAs for arginyl- and valyl-tRNA synthetases have the same molecular weight as the purified enzymes; translation of aspartyl-tRNA synthetase messenger RNA yielded a 68 kD molecular weight polypeptide (while the purified cristallisable enzyme appears as a 64-66 kD doublet, which, as we showed is a proteolysis product). The translation of the mRNAs for alpha and beta phenylalanyl-tRNA synthetase gave polypeptides having the same molecular weight as those obtained from the purified enzyme, but the major translation products are slightly heavier, indicating that they may be translated as precursors. As estimated from centrifugation experiments mRNAs of arginyl-, aspartyl-, alpha and beta subunits of phenylalanyl-tRNA synthetase were 1700-2000 nucleotides long, indicating that alpha and beta are translated from two different mRNAs. Images PMID:6344009

  12. Properties of 5-aminolaevulinate synthetase and its relationship to microsomal mixed-function oxidation in the southern armyworm (Spodoptera eridania).

    PubMed

    Brattsten, L B; Wilkinson, C F

    1975-07-01

    1. Activity of 5-aminolaevulinate synthetase was measured in the midgut and other tissues of the last larval instar of the southern armyworm (Spodoptera eridania Cramer, formerly Prodenia eridania Cramer). 2. Optimum conditions for measuring the activity were established with respect to all variables involved and considerable differences from those reported for mammalian enzyme preparations were found. 3. Maximum activity (20 nmol/h per mg of protein) occurs 18-24 h after the fifth moult and thereafter decreases to trace amounts as the larvae age and approach pupation. 4. Synthetase activity was rapidly induced by oral administration (in the diet) of pentamethylbenzene, phenobarbital, diethyl 1,4-dihydro-2,4,6-trimethylpyridine-3, 5-dicarboxylate, and 2-allyl-2-isopropylacetamide. 5. Puromycin inhibited the induction of synthetase by pentamethylbenzene. 6. Induction of 5-aminolaevulinate synthetase correlated well with the induction of microsomal N-demethylation of p-chloro-N-methylaniline, except for phenobarbital, which induced the microsomal oxidase relatively more than the synthetase. PMID:1004

  13. Properties of 5-aminolaevulinate synthetase and its relationship to microsomal mixed-function oxidation in the southern armyworm (Spodoptera eridania).

    PubMed Central

    Brattsten, L B; Wilkinson, C F

    1975-01-01

    1. Activity of 5-aminolaevulinate synthetase was measured in the midgut and other tissues of the last larval instar of the southern armyworm (Spodoptera eridania Cramer, formerly Prodenia eridania Cramer). 2. Optimum conditions for measuring the activity were established with respect to all variables involved and considerable differences from those reported for mammalian enzyme preparations were found. 3. Maximum activity (20 nmol/h per mg of protein) occurs 18-24 h after the fifth moult and thereafter decreases to trace amounts as the larvae age and approach pupation. 4. Synthetase activity was rapidly induced by oral administration (in the diet) of pentamethylbenzene, phenobarbital, diethyl 1,4-dihydro-2,4,6-trimethylpyridine-3, 5-dicarboxylate, and 2-allyl-2-isopropylacetamide. 5. Puromycin inhibited the induction of synthetase by pentamethylbenzene. 6. Induction of 5-aminolaevulinate synthetase correlated well with the induction of microsomal N-demethylation of p-chloro-N-methylaniline, except for phenobarbital, which induced the microsomal oxidase relatively more than the synthetase. PMID:1004

  14. Chitin synthetase activity is bound to chitosomes and to the plasma membrane in protoplasts of Saccharomyces cerevisiae.

    PubMed

    Flores Martinez, A; Schwencke, J

    1988-12-22

    The sub-cellular distribution of chitin synthetase was studied in homogenates of Saccharomyces cerevisiae protoplasts. Use of a mild disruption method minimized rupture of vacuoles and ensuing contamination of subcellular fractions by vacuolar proteinases. After fractionation of whole or partially purified homogenates through an isopycnic sucrose gradient chitin synthetase activity was found to be distributed between two distinct particulate fractions with different buoyant density and particle diameter. When whole homogenates were used, about 52% of the chitin synthetase loaded was localized in a microvesicular population identified as chitosomes (diameter 40-110 nm; buoyant density (d) = 1.146 g/cm3). Another vesicular population containing 26% of the activity was identified as plasma membrane vesicles because of its large mean diameter (260 nm), its high buoyant density (d = 1.203 g/cm3) and by the presence of the vanadate-sensitive ATPase activity. Moreover, after surface labeling of protoplasts with 3H-concanavalin A, the label cosedimented with the presumed plasma membrane vesicles. There was a negligible cross-contamination of the chitosome fraction by yeast plasma membrane markers. In both the plasma membrane and the chitosome fractions, the chitin synthetase was stable and essentially zymogenic. Activation of the chitosome fraction produces microfibrils 100-250 nm in length. Our results support the idea that chitosomes do not originate by plasma membrane vesiculation but are defined sub-cellular organelles containing most of the chitin synthetase in protoplasts of Saccharomyces cerevisiae. PMID:2974729

  15. Understanding species-specific differences in substrate recognition by Escherichia coli and human prolyl-tRNA synthetases.

    PubMed

    Musier-Forsyth, K; Stehlin, C; Burke, B; Liu, H

    1997-01-01

    Class II human prolyl-tRNA synthetase (ProRS) aminoacylates in vitro transcribed human tRNA(Pro) with kinetic parameters that are similar to those previously determined for aminoacylation of Escherichia coli tRNA(Pro) by its cognate synthetase. As in the bacterial system, large decreases in aminoacylation by human ProRS occur upon mutating anticodon positions G35 and G36 of human tRNA(Pro). The N73 'discriminator' base and the first and third base pairs of the acceptor stem vary between the E.coli and human isoacceptor groups. In contrast to the E. coli synthetase, the human enzyme does not appear to recognize these elements, since mutations at these positions do not significantly affect cognate synthetase charging. E. coli ProRS does not cross-aminoacylate human tRNA(Pro), and the bacterial tRNA(Pro) is a poor substrate for the human enzyme. Mutations in both the tRNAs and the synthetases have been made in an effort to identify elements in each system responsible for blocking cross-species aminoacylation. Alignment of all known ProRS primary sequences from different species reveals particularly low overall sequence homology, as well as two distinct groups of enzymes. The sequence divergence between E. coli and human ProRSs helps to explain the species-specific differences in the RNA code for aminoacylation of tRNA(Pro). PMID:9478190

  16. Compression of Laplacian pyramids through orthogonal transforms and improved prediction.

    PubMed

    Rath, Gagan; Yang, Wenxian; Guillemot, Christine

    2008-09-01

    Scalable representation of visual signals, such as image and video signals, has become a subject of active research since early 1980s. Scalability allows the adaptation of the bit rate and/or the resolution of the transmitted data to the network bandwidth and/or the rendering capability of the receiving device. For many years, spatial scalability has been achieved through wavelets, but recently the Laplacian pyramid (LP) has become an alternative choice because of reduced aliasing in the lower resolutions. In this paper, we focus on the coding efficiency of the LP with a view to transmitting it over a communication channel. In particular, we aim to improve the compression efficiency of the LP detail layers through improved interlayer prediction and orthogonal spatial transforms. First, we consider an LP in the open-loop configuration and propose to improve its rate-distortion performance by compressing it to a critically sampled representation. We derive four different orthogonal spatial transforms from the upsampling and downsampling filters that can achieve this representation, and apply them on the detail layers. The application of these transforms to the detail layers renders a fixed number of transform coefficients either zero or redundant, thus making their transmission unnecessary. Then we consider the compression of an LP in the closed-loop configuration through similar spatial transforms. Because of the introduction of quantization in the prediction loop, these spatial transforms applied on the detail layers do not produce the same number of zero or redundant transform coefficients as in the open-loop case. Nevertheless, the insight obtained from the open-loop coding leads us to enhance the interlayer prediction, and the subsequent application of the spatial transforms to the new detail layers aims to achieve better energy compaction. PMID:18701397

  17. Orthogonal Relations and Color Constancy in Dichromatic Colorblindness

    PubMed Central

    Pridmore, Ralph W.

    2014-01-01

    This paper employs uniform color space to analyze relations in dichromacy (protanopia, deuteranopia, tritanopia). Fifty percent or less of dichromats represent the classical reduction form of trichromacy, where one of three cones is inoperative but normal trichromatic color mixture such as complementary colors (pairs that mix white) are accepted by the dichromat, whose data can thus be plotted to CIE chromaticity spaces. The remaining dichromats comprise many and varied more-complex gene arrays from mutations, recombinations, etc. Though perhaps a minority, the three reductionist types provide a simple standard, in genotype and phenotype, to which the more complex remainder may be compared. Here, previously published data on dichromacy are plotted and analyzed in CIELUV uniform color space to find spatial relations in terms of color appearance space (e.g., hue angle). Traditional residual (seen) hues for protanopia and deuteranopia (both red–green colorblindness) are yellow and blue, but analysis indicates the protanopic residual hues are more greenish yellow and reddish blue than in tradition. Results for three illuminants (D65, D50, B) imply four principles in the spatial structure of dichromacy: (1) complementarity of confusion hue pairs and of residual hue pairs; (2) orthogonality of confusion locus and residual hues locus at their intersection with the white point, in each dichromatic type; (3) orthogonality of protanopic and tritanopic confusion loci; and (4) inverse relations between protanopic and tritanopic systems generally, such that one's confusion hues are the other's residual hues. Two of the three dichromatic systems do not represent components of normal trichromatic vision as sometimes thought but are quite different. Wavelength shifts between illuminants demonstrate chromatic adaptation correlates exactly with that in trichromatic vision. In theory these results clarify relations in and between types of dichromacy. They also apply in Munsell and

  18. Orthogonal arrays for computer experiments to assess important inputs

    SciTech Connect

    Moore, L. M.; McKay, Michael D.

    2002-01-01

    The topic of this paper is experiment planning, particularly fractional factorial designs or orthogonal arrays, for computer experiments to assess important inputs. The work presented in the paper is motivated by considering a non-stochastic computer simulation which has many inputs and which can, in a reasonable period of time, be run thousands of times. With many inputs, information that allows focus on a subset of important inputs is valuable. The characterization of 'importance' is expected to follow suggestions in McKay (1995) or McKay, et al. (1992). This analysis approach leads to considering factorial experiment designs. Inputs are associated with a finite number of discrete values, referred to as levels, so if each input has K levels and there are p inputs then there are K{sup P} possible distinct runs which constitute the K{sup P} factorial design space. The suggested size of p has been 35 to 50 so that even with K=2 the complete 2{sup P} factorial design space would not be run. Further, it is expected that the complexity of the simulation code and discrete levels possibly associated with equi-probable intervals from the input distribution make it desirable to consider more than 2 level inputs. Inputs levels of 5 and 7 have been investigated. In this paper, orthogonal array experiment designs, which are subsets of factorial designs also referred to as fractional factorial designs, are suggested as candidate experiments which provide meaningful basis for calculating and comparing R{sup 2} across subsets of inputs.

  19. Orthogonal relations and color constancy in dichromatic colorblindness.

    PubMed

    Pridmore, Ralph W

    2014-01-01

    This paper employs uniform color space to analyze relations in dichromacy (protanopia, deuteranopia, tritanopia). Fifty percent or less of dichromats represent the classical reduction form of trichromacy, where one of three cones is inoperative but normal trichromatic color mixture such as complementary colors (pairs that mix white) are accepted by the dichromat, whose data can thus be plotted to CIE chromaticity spaces. The remaining dichromats comprise many and varied more-complex gene arrays from mutations, recombinations, etc. Though perhaps a minority, the three reductionist types provide a simple standard, in genotype and phenotype, to which the more complex remainder may be compared. Here, previously published data on dichromacy are plotted and analyzed in CIELUV uniform color space to find spatial relations in terms of color appearance space (e.g., hue angle). Traditional residual (seen) hues for protanopia and deuteranopia (both red-green colorblindness) are yellow and blue, but analysis indicates the protanopic residual hues are more greenish yellow and reddish blue than in tradition. Results for three illuminants (D65, D50, B) imply four principles in the spatial structure of dichromacy: (1) complementarity of confusion hue pairs and of residual hue pairs; (2) orthogonality of confusion locus and residual hues locus at their intersection with the white point, in each dichromatic type; (3) orthogonality of protanopic and tritanopic confusion loci; and (4) inverse relations between protanopic and tritanopic systems generally, such that one's confusion hues are the other's residual hues. Two of the three dichromatic systems do not represent components of normal trichromatic vision as sometimes thought but are quite different. Wavelength shifts between illuminants demonstrate chromatic adaptation correlates exactly with that in trichromatic vision. In theory these results clarify relations in and between types of dichromacy. They also apply in Munsell and

  20. REGULATION OF RAT HEPATIC DELTA-AMINOLEVULINIC ACID SYNTHETASE AND HEME OXYGENASE ACTIVITIES: EVIDENCE FOR CONTROL BY HEME AND AGAINST MEDIATION BY PROSTHETIC IRON

    EPA Science Inventory

    The effects of in vivo administration of 6 compounds on the activity of delta-aminolevulinic acid (ALA) synthetase and heme oxygenase were determined. The order of decreasing potency in reducing ALA synthetase activity was heme, bilirubin, protoporphyrin IX, bilirubin dimethyl es...

  1. Recognition of Escherichia coli valine transfer RNA by its cognate synthetase: A fluorine-19 NMR study

    SciTech Connect

    Chu, Wenchy; Horowitz, J. )

    1991-02-12

    Interactions of 5-fluorouracil-substituted Escherichia coli tRNA{sup Val} with its cognate synthetase have been investigated by fluorine-19 nuclear magnetic resonance. Valyl-tRNA synthetase (VRS) (EC 6.1.1.9), purified to homogeneity from an overproducing strain of E. coli, differs somewhat from VRS previously isolated from E. coli K12. Its amino acid composition and N-terminal sequence agree well with results derived from the sequence of the VRS gene. Apparent K{sub M} and V{sub max} values of the purified VRS are the same for both normal and 5-fluorouracil (FUra)-substituted tRNA{sup Val}. Binding of VRS to (FUra)tRNA{sup Val} induces structural perturbations that are reflected in selective changes in the {sup 19}F NMR spectrum of the tRNA. Addition of increasing amounts of VRS results in a gradual loss of intensity at resonances corresponding to FU34, FU7, and FU67, with FU34, at the wobble position of the anticodon, being affected most. At higher VRS/tRNA ratios, a broadening and shifting of FU12 and of FU4 and/or FU8 occur. These results indicate that VRS interacts with tRNA{sup Val} along the entire inside of the L-shape molecule, from the acceptor stem to the anticodon. Valyl-tRNA synthetase also causes a splitting of resonances FU55 and FU64 in the T-loop and stem of tRNA{sup Val}, suggesting conformational changes in this part of the molecule. No {sup 19}F NMR evidence was found for formation of the Michael adduct between VRS and FU8 of 5-fluorouracil-substituted tRNA{sup Val} that has been proposed as a common intermediate in the aminoacylation reaction.

  2. Glial glutamate transporter and glutamine synthetase regulate GABAergic synaptic strength in the spinal dorsal horn.

    PubMed

    Jiang, Enshe; Yan, Xisheng; Weng, Han-Rong

    2012-05-01

    Decreased GABAergic synaptic strength ('disinhibition') in the spinal dorsal horn is a crucial mechanism contributing to the development and maintenance of pathological pain. However, mechanisms leading to disinhibition in the spinal dorsal horn remain elusive. We investigated the role of glial glutamate transporters (GLT-1 and GLAST) and glutamine synthetase in maintaining GABAergic synaptic activity in the spinal dorsal horn. Electrically evoked GABAergic inhibitory post-synaptic currents (eIPSCs), spontaneous IPSCs (sIPSCs) and miniature IPSCs were recorded in superficial spinal dorsal horn neurons of spinal slices from young adult rats. We used (2S,3S)-3-[3-[4-(trifluoromethyl)benzoylamino]benzyloxy]aspartate (TFB-TBOA), to block both GLT-1 and GLAST and dihydrokainic acid to block only GLT-1. We found that blockade of both GLAST and GLT-1 and blockade of only GLT-1 in the spinal dorsal horn decreased the amplitude of GABAergic eIPSCs, as well as both the amplitude and frequency of GABAergic sIPSCs or miniature IPSCs. Pharmacological inhibition of glial glutamine synthetase had similar effects on both GABAergic eIPSCs and sIPSCs. We provided evidence demonstrating that the reduction in GABAergic strength induced by the inhibition of glial glutamate transporters is due to insufficient GABA synthesis through the glutamate-glutamine cycle between astrocytes and neurons. Thus, our results indicate that deficient glial glutamate transporters and glutamine synthetase significantly attenuate GABAergic synaptic strength in the spinal dorsal horn, which may be a crucial synaptic mechanism underlying glial-neuronal interactions caused by dysfunctional astrocytes in pathological pain conditions. PMID:22339645

  3. Biochemical and Crystallographic Analysis of Substrate Binding and Conformational Changes in Acetyl-CoA Synthetase

    SciTech Connect

    Reger,A.; Carney, J.; Gulick, A.

    2007-01-01

    The adenylate-forming enzymes, including acyl-CoA synthetases, the adenylation domains of non-ribosomal peptide synthetases (NRPS), and firefly luciferase, perform two half-reactions in a ping-pong mechanism. We have proposed a domain alternation mechanism for these enzymes whereby, upon completion of the initial adenylation reaction, the C-terminal domain of these enzymes undergoes a 140{sup o} rotation to perform the second thioester-forming half-reaction. Structural and kinetic data of mutant enzymes support this hypothesis. We present here mutations to Salmonella enterica acetyl-CoA synthetase (Acs) and test the ability of the enzymes to catalyze the complete reaction and the adenylation half-reaction. Substitution of Lys609 with alanine results in an enzyme that is unable to catalyze the adenylate reaction, while the Gly524 to leucine substitution is unable to catalyze the complete reaction yet catalyzes the adenylation half-reaction with activity comparable to the wild-type enzyme. The positions of these two residues, which are located on the mobile C-terminal domain, strongly support the domain alternation hypothesis. We also present steady-state kinetic data of putative substrate-binding residues and demonstrate that no single residue plays a dominant role in dictating CoA binding. We have also created two mutations in the active site to alter the acyl substrate specificity. Finally, the crystallographic structures of wild-type Acs and mutants R194A, R584A, R584E, K609A, and V386A are presented to support the biochemical analysis.

  4. L-arginine recognition by yeast arginyl-tRNA synthetase.

    PubMed Central

    Cavarelli, J; Delagoutte, B; Eriani, G; Gangloff, J; Moras, D

    1998-01-01

    The crystal structure of arginyl-tRNA synthetase (ArgRS) from Saccharomyces cerevisiae, a class I aminoacyl-tRNA synthetase (aaRS), with L-arginine bound to the active site has been solved at 2.75 A resolution and refined to a crystallographic R-factor of 19.7%. ArgRS is composed predominantly of alpha-helices and can be divided into five domains, including the class I-specific active site. The N-terminal domain shows striking similarity to some completely unrelated proteins and defines a module which should participate in specific tRNA recognition. The C-terminal domain, which is the putative anticodon-binding module, displays an all-alpha-helix fold highly similar to that of Escherichia coli methionyl-tRNA synthetase. While ArgRS requires tRNAArg for the first step of the aminoacylation reaction, the results show that its presence is not a prerequisite for L-arginine binding. All H-bond-forming capability of L-arginine is used by the protein for the specific recognition. The guanidinium group forms two salt bridge interactions with two acidic residues, and one H-bond with a tyrosine residue; these three residues are strictly conserved in all ArgRS sequences. This tyrosine is also conserved in other class I aaRS active sites but plays several functional roles. The ArgRS structure allows the definition of a new framework for sequence alignments and subclass definition in class I aaRSs. PMID:9736621

  5. The McbB component of microcin B17 synthetase is a zinc metalloprotein.

    PubMed

    Zamble, D B; McClure, C P; Penner-Hahn, J E; Walsh, C T

    2000-12-26

    The microcin B17 synthetase converts glycine, serine, and cysteine residues in a polypeptide precursor into oxazoles and thiazoles during the maturation of the Escherichia coli antibiotic Microcin B17. This multimeric enzyme is composed of three subunits (McbB, McbC, and McbD), and it employs both ATP and FMN as cofactors. The McbB subunit was purified as a fusion with the maltose-binding protein (MBP), and metal analysis revealed that this protein binds 0.91+/-0.17 zinc atoms. Upon incubation of MBP-McbB with excess zinc, the stoichiometry increased to two atoms of zinc bound, but metal binding to the second site resulted in a decrease in the heterocyclization activity when MBP-McbB was reconstituted with the other components of the synthetase. Apo-protein was prepared by using p-hydroxymercuriphenylsulfonic acid (PMPS), and loss of the metal caused a severe reduction in enzymatic activity. However, if dithiothreitol was added to the PMPS reactions within a few minutes, enzymatic activity was retained and MBP-McbB could be reconstituted with zinc. Spectroscopic analysis of the cobalt-containing protein and extended X-ray absorption fine structure analysis of the zinc-containing protein both provide evidence for a tetrathiolate coordination sphere. Site-directed mutants of MBP-McbB as well as the synthetase tagged with the calmodulin-binding peptide were constructed. Activity assays and metal analysis were used to determine which of the six cysteines in McbB are metal ligands. These results suggest that the zinc cofactor in McbB plays a structural role. PMID:11123948

  6. Characterization of an Archaeal Medium-Chain Acyl Coenzyme A Synthetase from Methanosarcina acetivorans▿

    PubMed Central

    Meng, Yu; Ingram-Smith, Cheryl; Cooper, Leroy L.; Smith, Kerry S.

    2010-01-01

    Short- and medium-chain acyl coenzyme A (acyl-CoA) synthetases catalyze the formation of acyl-CoA from an acyl substrate, ATP, and CoA. These enzymes catalyze mechanistically similar two-step reactions that proceed through an enzyme-bound acyl-AMP intermediate. Here we describe the characterization of a member of this enzyme family from the methane-producing archaeon Methanosarcina acetivorans. This enzyme, a medium-chain acyl-CoA synthetase designated MacsMa, utilizes 2-methylbutyrate as its preferred substrate for acyl-CoA synthesis but cannot utilize acetate and thus cannot catalyze the first step of acetoclastic methanogenesis in M. acetivorans. When propionate or other less favorable acyl substrates, such as butyrate, 2-methylpropionate, or 2-methylvalerate, were utilized, the acyl-CoA was not produced or was produced at reduced levels. Instead, acyl-AMP and PPi were released in the absence of CoA, whereas in the presence of CoA, the intermediate was broken down into AMP and the acyl substrate, which were released along with PPi. These results suggest that although acyl-CoA synthetases may have the ability to utilize a broad range of substrates for the acyl-adenylate-forming first step of the reaction, the intermediate may not be suitable for the thioester-forming second step. The MacsMa structure has revealed the putative acyl substrate- and CoA-binding pockets. Six residues proposed to form the acyl substrate-binding pocket, Lys256, Cys298, Gly351, Trp259, Trp237, and Trp254, were targeted for alteration. Characterization of the enzyme variants indicates that these six residues are critical in acyl substrate binding and catalysis, and even conservative alterations significantly reduced the catalytic ability of the enzyme. PMID:20851904

  7. Affinity chromatography and affinity labeling of rat liver succinyl-CoA synthetase.

    PubMed

    Ball, D J; Nishimura, J S

    1980-11-25

    Succinyl-CoA synthetase has been purified to apparent homogeneity from rat liver. The key step in the purification procedure involved adsorption on a GDP dialdehyde (dial-GDP)-adipic dihydrazide-Sepharose 4B column and elution by GDP-Mg2+. Like the pig heart enzyme (Brownie, E. R., and Bridger, W. A. (1972) Can. J. Biochem. 50, 719--724), the rat liver enzyme was an alpha beta heterodimer and only the alpha subunit was phosphorylated by [gamma-32P]GTP. The A 280(0.1%) of the enzyme was determined to be 0.5. Amino acid analyses revealed significant similarities in 50% of the amino acid residues of rat liver and Escherichia coli succinyl-CoA synthetases. However, immunodiffusion analysis failed to reveal any antigenic identity between the two enzymes. Incubation with the affinity label, dial-GDP, in the presence of Mg2+ resulted in a biphasic inactivation of the enzyme. The extent of the rapid phase of inactivation appeared to be related to the extent of dephosphorylation of the enzyme and was prevented by preincubation of the enzyme with GTP-Mg2+. The presence of GDP-Mg2+ in the incubation medium prevented the slow phase of the inactivation and retarded the rapid phase. Dephosphorylated enzyme was approximately 2 orders of magnitude more susceptible to inactivation by dial-GDP than phosphorylated enzyme. Labeling of succinyl-CoA synthetase with [3H]dial-GDP gave a linear relationship between inactivation and incorporation of radioactivity with an extrapolated value of less than 1.2 mol of analog/mol of enzyme at 100% inactivation. The distribution of the label in enzyme that was inactivated 40% was approximately 60% in the alpha subunit and 40% in the beta subunit. Thus, while phosphorylation of the enzyme occurs exclusively in the alpha subunit, the nucleotide binding site appears to include components from both alpha and beta subunits. PMID:7430155

  8. Characterization of the acyl substrate binding pocket of acetyl-CoA synthetase.

    PubMed

    Ingram-Smith, Cheryl; Woods, Barrett I; Smith, Kerry S

    2006-09-26

    AMP-forming acetyl-CoA synthetase [ACS; acetate:CoA ligase (AMP-forming), EC 6.2.1.1] catalyzes the activation of acetate to acetyl-CoA in a two-step reaction. This enzyme is a member of the adenylate-forming enzyme superfamily that includes firefly luciferase, nonribosomal peptide synthetases, and acyl- and aryl-CoA synthetases/ligases. Although the structures of several superfamily members demonstrate that these enzymes have a similar fold and domain structure, the low sequence conservation and diversity of the substrates utilized have limited the utility of these structures in understanding substrate binding in more distantly related enzymes in this superfamily. The crystal structures of the Salmonella enterica ACS and Saccharomyces cerevisiae ACS1 have allowed a directed approach to investigating substrate binding and catalysis in ACS. In the S. enterica ACS structure, the propyl group of adenosine 5'-propylphosphate, which mimics the acyl-adenylate intermediate, lies in a hydrophobic pocket. Modeling of the Methanothermobacter thermautotrophicus Z245 ACS (MT-ACS1) on the S. cerevisiae ACS structure showed similar active site architecture, and alignment of the amino acid sequences of proven ACSs indicates that the four residues that compose the putative acetate binding pocket are well conserved. These four residues, Ile312, Thr313, Val388, and Trp416 of MT-ACS1, were targeted for alteration, and our results support that they do indeed form the acetate binding pocket and that alterations at these positions significantly alter the enzyme's affinity for acetate as well as the range of acyl substrates that can be utilized. In particular, Trp416 appears to be the primary determinant for acyl chain length that can be accommodated in the binding site. PMID:16981708

  9. Structural Insights into the Polyphyletic Origins of Glycyl tRNA Synthetases*♦

    PubMed Central

    Valencia-Sánchez, Marco Igor; Rodríguez-Hernández, Annia; Ferreira, Ruben; Santamaría-Suárez, Hugo Aníbal; Arciniega, Marcelino; Dock-Bregeon, Anne-Catherine; Moras, Dino; Beinsteiner, Brice; Brieba, Luis G.; Grøtli, Morten

    2016-01-01

    Glycyl tRNA synthetase (GlyRS) provides a unique case among class II aminoacyl tRNA synthetases, with two clearly widespread types of enzymes: a dimeric (α2) species present in some bacteria, archaea, and eukaryotes; and a heterotetrameric form (α2β2) present in most bacteria. Although the differences between both types of GlyRS at the anticodon binding domain level are evident, the extent and implications of the variations in the catalytic domain have not been described, and it is unclear whether the mechanism of amino acid recognition is also dissimilar. Here, we show that the α-subunit of the α2β2 GlyRS from the bacterium Aquifex aeolicus is able to perform the first step of the aminoacylation reaction, which involves the activation of the amino acid with ATP. The crystal structure of the α-subunit in the complex with an analog of glycyl adenylate at 2.8 Å resolution presents a conformational arrangement that properly positions the cognate amino acid. This work shows that glycine is recognized by a subset of different residues in the two types of GlyRS. A structural and sequence analysis of class II catalytic domains shows that bacterial GlyRS is closely related to alanyl tRNA synthetase, which led us to define a new subclassification of these ancient enzymes and to propose an evolutionary path of α2β2 GlyRS, convergent with α2 GlyRS and divergent from AlaRS, thus providing a possible explanation for the puzzling existence of two proteins sharing the same fold and function but not a common ancestor. PMID:27226617

  10. Molecular cloning and characterization of an S-adenosylmethionine synthetase gene from Chorispora bungeana.

    PubMed

    Ding, Chenchen; Chen, Tao; Yang, Yu; Liu, Sha; Yan, Kan; Yue, Xiule; Zhang, Hua; Xiang, Yun; An, Lizhe; Chen, Shuyan

    2015-11-10

    S-adenosylmethionine synthetase (SAMS) catalyzes the formation of S-adenosylmethionine (SAM) which is a molecule essential for polyamines and ethylene biosynthesis, methylation modifications of protein, DNA and lipids. SAMS also plays an important role in abiotic stress response. Chorispora bungeana (C. bungeana) is an alpine subnival plant species which possesses strong tolerance to cold stress. Here, we cloned and characterized an S-adenosylmethionine synthetase gene, CbSAMS (C. bungeana S-adenosylmethionine synthetase), from C. bungeana, which encodes a protein of 393 amino acids containing a methionine binding motif GHPDK, an ATP binding motif GAGDQG and a phosphate binding motif GGGAFSGDK. Furthermore, an NES (nuclear export signal) peptide was identified through bioinformatics analysis. To explore the CbSAMS gene expression regulation, we isolated the promoter region of CbSAMS gene 1919bp upstream the ATG start codon, CbSAMSp, and analyzed its cis-acting elements by bioinformatics method. It was revealed that a transcription start site located at 320 bp upstream the ATG start codon and cis-acting elements related to light, ABA, auxin, ethylene, MeJA, low temperature and drought had been found in the CbSAMSp sequence. The gene expression pattern of CbSAMS was then analyzed by TR-qPCR and GUS assay method. The result showed that CbSAMS is expressed in all examined tissues including callus, roots, petioles, leaves, and flowers with a significant higher expression level in roots and flowers. Furthermore, the expression level of CbSAMS was induced by low temperature, ethylene and NaCl. Subcellular localization revealed that CbSAMS was located in the cytoplasm and nucleus but has a significant higher level in the nucleus. These results indicated a potential role of CbSAMS in abiotic stresses and plant growth in C. bungeana. PMID:26205258

  11. Bacterial expression of catalytically active fragments of the multifunctional enzyme enniatin synthetase.

    PubMed

    Haese, A; Pieper, R; von Ostrowski, T; Zocher, R

    1994-10-14

    Enniatin synthetase catalyzes the biosynthesis of N-methylated cyclohexadepsipeptides. The 347 kDa enzyme is encoded by the esyn1 gene of Fusarium scirpi and contains two domains (EA and EB) homologous to each other and to regions of other microbial peptide synthetases. Parts of the esyn1 gene were subcloned in frame to a small lacZ gene portion of Escherichia coli expression vectors. Overproduced recombinant proteins showed a high tendency towards inclusion body formation and could be only partially dissolved in 8 M urea or 6 M guanidine hydrochloride. After renaturation, a 121 kDa recombinant protein representing the N-terminal conserved domain EA of enniatin synthetase was shown to activate D-hydroxyisolvaleric acid via adenylation. Similarly, a 158 kDa recombinant protein comprising the C-terminal conserved domain EB catalyzed the activation of the substrate amino acid (e.g. L-valine). Moreover, this protein could be photolabeled with S-[methyl-14C]adenosyl-L-methionine, (AdoMet) indicating the presence of the methyltransferase. Both functions, L-valine activation and AdoMet binding, could be assigned to a 108 kDa recombinant protein encompassing the A and the M segment of domain EB. The fact that a 65 kDa recombinant protein representing the M portion could be photolabeled, indicated the localization of the methyltransferase in this region. Three deletion mutants of the 65 kDa protein were shown to be inactive with respect to UV-induced AdoMet labeling. PMID:7932733

  12. Cylindrospermopsin and Saxitoxin Synthetase Genes in Cylindrospermopsis raciborskii Strains from Brazilian Freshwater

    PubMed Central

    Hoff-Risseti, Caroline; Dörr, Felipe Augusto; Schaker, Patricia Dayane Carvalho; Pinto, Ernani; Werner, Vera Regina; Fiore, Marli Fatima

    2013-01-01

    The Cylindrospermopsis raciborskii population from Brazilian freshwater is known to produce saxitoxin derivatives (STX), while cylindrospermopsin (CYN), which is commonly detected in isolates from Australia and Asia continents, has thus far not been detected in South American strains. However, during the investigation for the presence of cyrA, cyrB, cyrC and cyrJ CYN synthetase genes in the genomes of four laboratory-cultured C. raciborskii Brazilian strains, the almost complete cyrA gene sequences were obtained for all strains, while cyrB and cyrC gene fragments were observed in two strains. These nucleotide sequences were translated into amino acids, and the predicted protein functions and domains confirmed their identity as CYN synthetase genes. Attempts to PCR amplify cyrJ gene fragments from the four strains were unsuccessful. Phylogenetic analysis grouped the nucleotide sequences together with their homologues found in known CYN synthetase clusters of C. raciborskii strains with high bootstrap support. In addition, fragments of sxtA, sxtB and sxtI genes involved in STX production were also obtained. Extensive LC-MS analyses were unable to detect CYN in the cultured strains, whereas the production of STX and its analogues was confirmed in CENA302, CENA305 and T3. To our knowledge, this is the first study reporting the presence of cyr genes in South American strains of C. raciborskii and the presence of sxt and cyr genes in a single C. raciborskii strain. This discovery suggests a shift in the type of cyanotoxin production over time of South American strains of C. raciborskii and contributes to the reconstruction of the evolutionary history and diversification of cyanobacterial toxins. PMID:24015317

  13. Control of Escherichia coli lysyl-tRNA synthetase expression by anaerobiosis.

    PubMed Central

    Lévêque, F; Gazeau, M; Fromant, M; Blanquet, S; Plateau, P

    1991-01-01

    Escherichia coli lysyl-tRNA synthetase was previously shown to occur as two distinct species encoded by either the lysS or the lysU gene. The expression of one of these genes, lysU, is under the control of cell growth conditions. To study the regulation of lysU, delta lysS strains were constructed. During aerobic growth at 37 degrees C or below, the amount of the lysU product in the cell is so reduced that delta lysS bacteria grow only poorly. The reduced expression of lysU is not related to the steady-state lysyl-tRNA synthetase concentration in the cell, since the expression of a lysU::lacZ fusion is insensitive to the absence of either lysS or lysU or to the addition of a multi-copy plasmid carrying either lysU or lysS. During anaerobic growth in rich medium, the lysU gene becomes strongly expressed and, in cell extracts, the amount of lysyl-tRNA synthetase activity originating from lysU may become seven times greater than the activity originating from lysS. In minimal medium, lysU expression is only slightly induced. Evidence that the sensitivity of lysU expression to anaerobiosis, as well as to low external pH conditions (E. W. Hickey and I. N. Hirshfield, Appl. Environ. Microbiol. 56:1038-1045, 1990), is governed at the level of transcription is provided. Images FIG. 4 PMID:1744045

  14. Structural Insights into the Polyphyletic Origins of Glycyl tRNA Synthetases.

    PubMed

    Valencia-Sánchez, Marco Igor; Rodríguez-Hernández, Annia; Ferreira, Ruben; Santamaría-Suárez, Hugo Aníbal; Arciniega, Marcelino; Dock-Bregeon, Anne-Catherine; Moras, Dino; Beinsteiner, Brice; Mertens, Haydyn; Svergun, Dmitri; Brieba, Luis G; Grøtli, Morten; Torres-Larios, Alfredo

    2016-07-01

    Glycyl tRNA synthetase (GlyRS) provides a unique case among class II aminoacyl tRNA synthetases, with two clearly widespread types of enzymes: a dimeric (α2) species present in some bacteria, archaea, and eukaryotes; and a heterotetrameric form (α2β2) present in most bacteria. Although the differences between both types of GlyRS at the anticodon binding domain level are evident, the extent and implications of the variations in the catalytic domain have not been described, and it is unclear whether the mechanism of amino acid recognition is also dissimilar. Here, we show that the α-subunit of the α2β2 GlyRS from the bacterium Aquifex aeolicus is able to perform the first step of the aminoacylation reaction, which involves the activation of the amino acid with ATP. The crystal structure of the α-subunit in the complex with an analog of glycyl adenylate at 2.8 Å resolution presents a conformational arrangement that properly positions the cognate amino acid. This work shows that glycine is recognized by a subset of different residues in the two types of GlyRS. A structural and sequence analysis of class II catalytic domains shows that bacterial GlyRS is closely related to alanyl tRNA synthetase, which led us to define a new subclassification of these ancient enzymes and to propose an evolutionary path of α2β2 GlyRS, convergent with α2 GlyRS and divergent from AlaRS, thus providing a possible explanation for the puzzling existence of two proteins sharing the same fold and function but not a common ancestor. PMID:27226617

  15. Glutamine synthetase in ribulose 1,5-bisphosphate carboxylase/oxygenase deficient tobacco mutants in cell suspension culture.

    PubMed

    Hirel, B; Nato, A; Martin, F

    1984-06-01

    In two tobacco mutants lacking ribulose, 1,5-bisphosphate carboxylase/oxygenase the amount of glutamine synthetase and its activity were determined and compared with the wild type green cells. It was shown that in these two mutants glutamine synthetase protein content was six times lower than in the wild type. This situation was comparable to that found in etiolated cells where ribulose 1,5-bisphosphate carboxylase/oxygenase was absent. These observations suggest that a common regulatory mechanism might control the dual light dependent biosynthesis of both enzymes. The results have also implications concerning the efficiency of the reassimilation of ammonia by chloroplastic glutamine synthetase during the photorespiratory process. PMID:24253436

  16. Purification, gene cloning, and characterization of γ-butyrobetainyl CoA synthetase from Agrobacterium sp. 525a.

    PubMed

    Fujimitsu, Hiroshi; Matsumoto, Akira; Takubo, Sayaka; Fukui, Akiko; Okada, Kazuma; Mohamed Ahmed, Isam A; Arima, Jiro; Mori, Nobuhiro

    2016-08-01

    The report is the first of purification, overproduction, and characterization of a unique γ-butyrobetainyl CoA synthetase from soil-isolated Agrobacterium sp. 525a. The primary structure of the enzyme shares 70-95% identity with those of ATP-dependent microbial acyl-CoA synthetases of the Rhizobiaceae family. As distinctive characteristics of the enzyme of this study, ADP was released in the catalytic reaction process, whereas many acyl CoA synthetases are annotated as an AMP-forming enzyme. The apparent Km values for γ-butyrobetaine, CoA, and ATP were, respectively, 0.69, 0.02, and 0.24 mM. PMID:27125317

  17. Alterations in Dihydropteroate Synthetase in Cell-Free Extracts of Sulfanilamide-Resistant Neisseria meningitidis and Neisseria gonorrhoeae

    PubMed Central

    Ho, Richard I.; Corman, Leonard; Morse, Stephen A.; Artenstein, Malcolm S.

    1974-01-01

    Extracts from Neisseria meningitidis and N. gonorrhoeae with varying susceptibility to sulfanilamide have been investigated for dihydropteroate synthetase activity. Sulfanilamide was a competitive inhibitor of dihydropteroate synthetase with respect to p-aminobenzoate in extracts from both species. Though the Km for p-aminobenzoate was unaffected, the Ki for sulfanilamide increased and the Vmax decreased as the strains' resistance to sulfanilamide increased. Temperature studies have revealed differences in the dihydropteroate synthetase from N. meningitidis and N. gonorrhoeae. A direct relationship was observed between the minimal inhibitory concentration of sulfanilamide determined in vitro and the ratio of Ki/Km. This ratio may be a molecular explanation of sulfanilamide resistance for both N. meningitidis and N. gonorrhoeae. PMID:15825393

  18. Inhibition of human glutamine synthetase by L-methionine-S,R-sulfoximine-relevance to the treatment of neurological diseases.

    PubMed

    Jeitner, Thomas M; Cooper, Arthur J L

    2014-12-01

    At high concentrations, the glutamine synthetase inhibitor L-methionine-S,R-sulfoximine (MSO) is a convulsant, especially in dogs. Nevertheless, sub-convulsive doses of MSO are neuroprotective in rodent models of hyperammonemia, acute liver disease, and amyotrophic lateral sclerosis and suggest MSO may be clinically useful. Previous work has also shown that much lower doses of MSO are required to produce convulsions in dogs than in primates. Evidence from the mid-20th century suggests that humans are also less sensitive. In the present work, the inhibition of recombinant human glutamine synthetase by MSO is shown to be biphasic-an initial reversible competitive inhibition (K i 1.19 mM) is followed by rapid irreversible inactivation. This K i value for the human enzyme accounts, in part, for relative insensitivity of primates to MSO and suggests that this inhibitor could be used to safely inhibit glutamine synthetase activity in humans. PMID:24136581

  19. Inhibition of human glutamine synthetase by L-methionine-S,R-sulfoximine – relevance to the treatment of neurological diseases

    PubMed Central

    Jeitner, Thomas M.; Cooper, Arthur J. L.

    2013-01-01

    At high concentrations, the glutamine synthetase inhibitor L-methionine-S,R-sulfoximine is a convulsant, especially in dogs. Nevertheless, sub-convulsive doses of MSO are neuroprotective in rodent models of hyperammonemia, acute liver disease, and amyotrophic lateral sclerosis and suggest MSO may be clinically useful. Previous work has also shown that much lower doses of MSO are required to produce convulsions in dogs than in primates. Evidence from the mid-20th century suggests that humans are also less sensitive. In the present work, the inhibition of recombinant human glutamine synthetase with MSO is shown to be biphasic – an initial reversible competitive inhibition (Ki 1.19 mM) is followed by rapid irreversible inactivation. This Ki value for the human enzyme accounts, in part, for relative insensitivity of primates to MSO and suggests that this inhibitor could be used to safely inhibit glutamine synthetase activity in humans. PMID:24136581

  20. Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of DHNA synthetase from Geobacillus kaustophilus

    SciTech Connect

    Kanaujia, Shankar Prasad; Ranjani, Chellamuthu Vasuki; Jeyakanthan, Jeyaraman; Baba, Seiki; Kuroishi, Chizu; Ebihara, Akio; Shinkai, Akeo; Kuramitsu, Seiki; Shiro, Yoshitsugu; Sekar, Kanagaraj; Yokoyama, Shigeyuki

    2007-02-01

    DHNA synthetase from G. kaustophilus has been cloned, expressed, purified and crystallized. The aerobic Gram-positive bacterium Geobacillus kaustophilus is a bacillus species that was isolated from deep-sea sediment from the Mariana Trench. 1,4-Dihydroxy-2-naphthoate (DHNA) synthetase plays a vital role in the biosynthesis of menaquinone (vitamin K{sub 2}) in this bacterium. DHNA synthetase from Geobacillus kaustophilus was crystallized in the orthorhombic space group C222{sub 1}, with unit-cell parameters a = 77.01, b = 130.66, c = 131.69 Å. The crystal diffracted to a resolution of 2.2 Å. Preliminary studies and molecular-replacement calculations reveal the presence of three monomers in the asymmetric unit.

  1. Hydrogen peroxide induces spawning in mollusks, with activation of prostaglandin endoperoxide synthetase.

    PubMed

    Morse, D E; Duncan, H; Hooker, N; Morse, A

    1977-04-15

    Addition of hydrogen peroxide to seawater causes synchronous spawning in gravid male and female abalones, and certain other mollusks as well. This effect is blocked by exposure of the animals to aspirin, an inhibitor of the enzyme catalyzing oxidative synthesis of prostaglandin endoperoxide. Hydrogen peroxide activates this enzymatic reaction in cell-free extracts prepared from abalone eggs (a very rich source of the prostaglandin endoperoxide synthetase); this effect appears to reveal a fundamental property of prostaglandin endoperoxide synthesis. Applicability of these findings to both mariculture and medical purposes is suggested. PMID:403609

  2. Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. The involvement of sulphur metabolism

    PubMed Central

    Neuberger, Albert; Sandy, John D.; Tait, George H.

    1973-01-01

    1. The `initial' 5-aminolaevulinate synthetase activity, that is the activity observed immediately after cell disruption, in extracts prepared from unharvested semianaerobically grown Rhodopseudomonas spheroides, was twice that observed under the same assay conditions in extracts prepared from harvested cells. 2. The effect of oxygenation of a culture on the `maximum' aminolaevulinate synthetase activity, that is the activity observed 1h after disruption of harvested cells, is markedly influenced by the contents of the growth medium. Oxygenation of organisms for 1h in the medium in which they have grown produces an 80–90% decrease in maximum activity, whereas similar treatment of organisms resuspended in fresh medium produces less than a 40% decrease. 3. This protective effect of fresh medium is absolutely dependent on the presence of sulphate. When cells are suspended in sulphate-deficient fresh medium, the maximum activity falls by 65–75% even without oxygenation. A high maximum activity is regenerated when sulphate is resupplied. 4. When organisms are oxygenated in the medium in which they have grown, the cellular contents of GSH+GSSG and cysteine+cystine fall very markedly and homolanthionine is formed. Both the fall in aminolaevulinate synthetase activity and the changes in sulphur metabolism are largely prevented by the addition of compounds which stimulate synthesis of cysteine de novo or inhibit the conversion of cysteine S into homocysteine S. 5. The maximum aminolaevulinate synthetase activity was directly proportional to the GSH+GSSG content of all cell preparations. In glutathione-depleted extracts the `low'-activity enzyme could be re-activated in vitro by the addition of GSH, GSSG, cysteine or cystine, whereas in extracts with a high glutathione content the `high'-activity enzyme was unaffected by these sulphur compounds. 6. The activation of low-activity enzyme with exogenous sulphur compounds was prevented by excluding air or by adding NADH

  3. Argininosuccinate synthetase as a plasma biomarker of liver injury after acetaminophen overdose in rodents and humans

    PubMed Central

    McGill, Mitchell R.; Cao, Mengde; Svetlov, Archie; Sharpe, Matthew R.; Williams, C. David; Curry, Steven C.; Farhood, Anwar; Jaeschke, Hartmut; Svetlov, Stanislav I.

    2014-01-01

    Context New biomarkers are needed in acetaminophen (APAP) hepatotoxicity. Plasma argininosuccinate synthetase (ASS) is a promising candidate. Objective Characterize ASS in APAP hepatotoxicity. Methods ASS was measured in plasma from rodents and humans with APAP hepatotoxicity. Results In mice, ASS increased before injury, peaked before ALT, and decreased rapidly. Fischer rats had a greater increase in ASS relative to ALT. Patients with abnormal liver test results had very high ASS compared to controls. ASS appeared to increase early in some patients, and declined rapidly in all. Conclusions : ASS may be a useful biomarker of acute cell death in APAP hepatotoxicity. PMID:24597531

  4. Overexpression of acetyl-CoA synthetase in Saccharomyces cerevisiae increases acetic acid tolerance

    PubMed Central

    Ding, Jun; Holzwarth, Garrett; Penner, Michael H.; Patton-Vogt, Jana; Bakalinsky, Alan T.

    2015-01-01

    Acetic acid-mediated inhibition of the fermentation of lignocellulose-derived sugars impedes development of plant biomass as a source of renewable ethanol. In order to overcome this inhibition, the capacity of Saccharomyces cerevisiae to synthesize acetyl-CoA from acetic acid was increased by overexpressing ACS2 encoding acetyl-coenzyme A synthetase. Overexpression of ACS2 resulted in higher resistance to acetic acid as measured by an increased growth rate and shorter lag phase relative to a wild-type control strain, suggesting that Acs2-mediated consumption of acetic acid during fermentation contributes to acetic acid detoxification. PMID:25673654

  5. Tissue Distribution of Glutamate Synthase and Glutamine Synthetase in Rice Leaves 1

    PubMed Central

    Yamaya, Tomoyuki; Hayakawa, Toshihiko; Tanasawa, Keisuke; Kamachi, Kazunari; Mae, Tadahiko; Ojima, Kunihiko

    1992-01-01

    To further explore the function of NADH-dependent glutamate synthase (GOGAT), the tissue distribution of NADH-GOGAT protein and activity was investigated in rice (Oryza sativa L.) leaves. The distributions of ferredoxin (Fd)-dependent GOGAT, plastidic glutamine synthetase, and cytosolic glutamine synthetase proteins were also determined in the same tissues. High levels of NADH-GOGAT protein (33.1 μg protein/g fresh weight) and activity were detected in the 10th leaf blade before emergence. The unexpanded, nongreen portion of the 9th leaf blade contained more than 50% of the NADH-GOGAT protein and activity per gram fresh weight when compared with the 10th leaf. The expanding, green portion of the 9th leaf blade outside of the sheath contained a slightly lower abundance of NADH-GOGAT protein than the nongreen portion of the 9th blade on a fresh weight basis. The fully expanded leaf blades at positions lower than the 9th leaf had decreased NADH-GOGAT levels as a function of increasing age, and the oldest, 5th blade contained only 4% of the NADH-GOGAT protein compared with the youngest 10th leaf blade. Fd-GOGAT protein, on the other hand, was the major form of GOGAT in the green tissues, and the highest amount of Fd-GOGAT protein (111 μg protein/g fresh weight) was detected in the 7th leaf blade. In the nongreen 10th leaf blade, the content of Fd-GOGAT protein was approximately 7% of that found in the 7th leaf blade. In addition, the content of NADH-GOGAT protein in the 10th leaf blade was about 4 times higher than that of Fd-GOGAT protein. The content of plastidic glutamine synthetase polypeptide was also the highest in the 7th leaf blade (429 μg/g fresh weight) and lowest in nongreen blades and sheaths. On the other hand, the relative abundance of the cytosolic glutamine synthetase polypeptide was the highest in the oldest leaf blade, decreasing to 10 to 20% of that value in young, nongreen leaves. These results suggest that NADH-GOGAT is important for the

  6. The Role of Pyruvate Dehydrogenase and Acetyl-Coenzyme A Synthetase in Fatty Acid Synthesis in Developing Arabidopsis Seeds1

    PubMed Central

    Ke, Jinshan; Behal, Robert H.; Back, Stephanie L.; Nikolau, Basil J.; Wurtele, Eve Syrkin; Oliver, David J.

    2000-01-01

    Acetyl-coenzyme A (acetyl-CoA) formed within the plastid is the precursor for the biosynthesis of fatty acids and, through them, a range of important biomolecules. The source of acetyl-CoA in the plastid is not known, but two enzymes are thought to be involved: acetyl-CoA synthetase and plastidic pyruvate dehydrogenase. To determine the importance of these two enzymes in synthesizing acetyl-CoA during lipid accumulation in developing Arabidopsis seeds, we isolated cDNA clones for acetyl-CoA synthetase and for the ptE1α- and ptE1β-subunits of plastidic pyruvate dehydrogenase. To our knowledge, this is the first reported acetyl-CoA synthetase sequence from a plant source. The Arabidopsis acetyl-CoA synthetase preprotein has a calculated mass of 76,678 D, an apparent plastid targeting sequence, and the mature protein is a monomer of 70 to 72 kD. During silique development, the spatial and temporal patterns of the ptE1β mRNA level are very similar to those of the mRNAs for the plastidic heteromeric acetyl-CoA carboxylase subunits. The pattern of ptE1β mRNA accumulation strongly correlates with the formation of lipid within the developing embryo. In contrast, the level of mRNA for acetyl-CoA synthetase does not correlate in time and space with lipid accumulation. The highest level of accumulation of the mRNA for acetyl-CoA synthetase during silique development is within the funiculus. These mRNA data suggest a predominant role for plastidic pyruvate dehydrogenase in acetyl-CoA formation during lipid synthesis in seeds. PMID:10859180

  7. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture

    SciTech Connect

    Torreira, Eva; Seabra, Ana Rita; Marriott, Hazel; Zhou, Min; Llorca, Óscar; Robinson, Carol V.; Carvalho, Helena G.; Fernández-Tornero, Carlos; Pereira, Pedro José Barbosa

    2014-04-01

    The experimental models of dicotyledonous cytoplasmic and plastid-located glutamine synthetases unveil a conserved eukaryotic-type decameric architecture, with subtle structural differences in M. truncatula isoenzymes that account for their distinct herbicide resistance. The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants.

  8. 2D nearly orthogonal mesh generation with controls on distortion functions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A method to control the distortion function of the Ryskin and Leal (RL) orthogonal mesh generation system is presented. The proposed method considers the effects from not only the local orthogonal condition but also the local smoothness condition (the geometry and the mesh size) on the distortion fu...

  9. A note on the zeros of Freud-Sobolev orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Moreno-Balcazar, Juan J.

    2007-10-01

    We prove that the zeros of a certain family of Sobolev orthogonal polynomials involving the Freud weight function e-x4 on are real, simple, and interlace with the zeros of the Freud polynomials, i.e., those polynomials orthogonal with respect to the weight function e-x4. Some numerical examples are shown.

  10. Families of orthogonal Schrödinger cat-like-states

    NASA Astrophysics Data System (ADS)

    Praxmeyer, Ludmiła

    2016-06-01

    We analyze the condition of orthogonality between optical Schrödinger cat-like-states constructed as a superposition of two coherent states. We show that the orthogonality condition leads to the quantization of values of a naturally emerging symplectic form, while values of the corresponding metric form are continuous. A complete analytical solution of the problem is presented.

  11. A pillar[5]arene-fused cryptand: from orthogonal self-assembly to supramolecular polymer.

    PubMed

    Wang, Qi; Cheng, Ming; Zhao, Yue; Wu, Lin; Jiang, Juli; Wang, Leyong; Pan, Yi

    2015-02-28

    A pillar[5]arene-fused cryptand with two different cavities was synthesized successfully. It was found that the novel tricyclic host could associate with two different guest species in an orthogonal manner. And based on this orthogonal self-assembly of two host-guest interactions, a novel type of supramolecular polymer was constructed easily and conveniently. PMID:25636005

  12. Molecular recognition of proline tRNA by prolyl-tRNA synthetase from hyperthermophilic archaeon, Aeropyrum pernix K1.

    PubMed

    Yokozawa, Junji; Okamoto, Koji; Kawarabayasi, Yutaka; Kuno, Atsushi; Hasegawa, Tsunemi

    2003-01-01

    To investigate the recognition mechanism of tRNA(Pro) by prolyl-tRNA synthetase from hyperthermophilic archaeon, Aeropyrum pernix K1, various tRNA(Pro) transcripts were prepared by in vitro transcription system. These transcripts were aminoacylated with proline by overexpressed A. pernix prolyl-tRNA synthetase. From prolylation experiments, recognition elements of A. pernix tRNA(Pro) were determined to be G35 and G36 of anticodon, discriminator base A73, and G1-C72 base pair at acceptor stem end. PMID:14510473

  13. Activity of interferon-dependent 2',5'-oligoadenylate synthetase in rat lymphoid cells under transformed environment conditions

    NASA Astrophysics Data System (ADS)

    Ostapchenko, L. I.; Mikhailik, I. V.; Prokopova, K. V.

    It is detected that interferon-dependent 2',5'-oligoadenylate synthetase is a sensitive index of immunocompetent cells functional state under transformed environment conditions. Microgravitation and ionising radiation induce increase of investigated enzyme activity in rat lymphocytes, which can be a result of lymphoid cells compensatory mechanisms starting in response to stress factors action. Administration of interferon inductors permits to stimulate the 2',5'-oligoadenylate synthetase, which enables one to correct pathological changes in the cells and to intensify adaptive reactions of immune systems.

  14. The nucleotide sequence of the promoter region of hisS, the structural gene for histidyl-tRNA synthetase.

    PubMed

    Eisenbeis, S J; Parker, J

    1982-05-01

    A plasmid has been constructed which carries hisS, the structural gene for histidyl-RNA synthetase of E. coli, on a 1.6-kb fragment bounded by PvuII and BstEII sites. The DNA sequence of both ends of this fragment was determined. The amino-terminal sequence of histidyl-tRNA synthetase was also determined to locate the promoter proximal coding region and the frame in which it is read. Three promoters were identified by consensus criteria. The region surrounding these promoters contains extensive twofold symmetry. PMID:6290315

  15. Prostaglandin endoperoxide synthetase and the activation of benzo(a)pyrene to reactive metabolites in vivo in guinea pigs

    SciTech Connect

    Garattini, E.; Coccia, P.; Romano, M.; Jiritano, L.; Noseda, A.; Salmona, M.

    1984-11-01

    The role of prostaglandin endoperoxide synthetase in the in vivo activation of benzo(a)pyrene to reactive metabolites capable of interacting irreversibly with cellular macromolecules was studied in guinea pig liver, lung, kidney, spleen, small intestine, colon, and brain. DNA and protein covalent binding experiments were made after systemic administration of acetylsalicylic acid (200 mg/kg) followed by radiolabeled benzo(a)pyrene (4 microgram/kg). Results are compared with a control situation in which the prostaglandin endoperoxide synthetase inhibitor (acetylsalicylic acid) was not administered. No decrease in the level of DNA or protein benzo(a)pyrene-derived covalent binding was observed in any of the tissues studied.

  16. Orthogonal analysis of functional gold nanoparticles for biomedical applications.

    PubMed

    Tsai, De-Hao; Lu, Yi-Fu; DelRio, Frank W; Cho, Tae Joon; Guha, Suvajyoti; Zachariah, Michael R; Zhang, Fan; Allen, Andrew; Hackley, Vincent A

    2015-11-01

    We report a comprehensive strategy based on implementation of orthogonal measurement techniques to provide critical and verifiable material characteristics for functionalized gold nanoparticles (AuNPs) used in biomedical applications. Samples were analyzed before and after ≈50 months of cold storage (≈4 °C). Biomedical applications require long-term storage at cold temperatures, which could have an impact on AuNP therapeutics. Thiolated polyethylene glycol (SH-PEG)-conjugated AuNPs with different terminal groups (methyl-, carboxylic-, and amine-) were chosen as a model system due to their high relevancy in biomedical applications. Electrospray-differential mobility analysis, asymmetric-flow field flow fractionation, transmission electron microscopy, scanning electron microscopy, atomic force microscopy, inductively coupled plasma mass spectrometry, and small-angle X-ray scattering were employed to provide both complementary and orthogonal information on (1) particle size and size distribution, (2) particle concentrations, (3) molecular conjugation properties (i.e., conformation and surface packing density), and (4) colloidal stability. Results show that SH-PEGs were conjugated on the surface of AuNPs to form a brush-like polymer corona. The surface packing density of SH-PEG was ≈0.42 nm(-2) for the methyl-PEG-SH AuNPs, ≈0.26 nm(-2) for the amine-SH-PEG AuNPs, and ≈0.18 nm(-2) for the carboxylic-PEG-SH AuNPs before cold storage, approximately 10 % of its theoretical maximum value. The conformation of surface-bound SH-PEGs was then estimated to be in an intermediate state between brush-like and random-coiled, based on the measured thicknesses in liquid and in dry states. By analyzing the change in particle size distribution and number concentration in suspension following cold storage, the long term colloidal stability of AuNPs was shown to be significantly improved via functionalization with SH-PEG, especially in the case of methyl-PEG-SH and carboxylic

  17. Anticodon recognition in evolution: switching tRNA specificity of an aminoacyl-tRNA synthetase by site-directed peptide transplantation.

    PubMed

    Brevet, Annie; Chen, Josiane; Commans, Stéphane; Lazennec, Christine; Blanquet, Sylvain; Plateau, Pierre

    2003-08-15

    The highly conserved aspartyl-, asparaginyl-, and lysyl-tRNA synthetases compose one subclass of aminoacyl-tRNA synthetases, called IIb. The three enzymes possess an OB-folded extension at their N terminus. The function of this extension is to specifically recognize the anticodon triplet of the tRNA. Three-dimensional models of bacterial aspartyl- and lysyl-tRNA synthetases complexed to tRNA indicate that a rigid scaffold of amino acid residues along the five beta-strands of the OB-fold accommodates the base U at the center of the anticodon. The binding of the adjacent anticodon bases occurs through interactions with a flexible loop joining strands 4 and 5 (L45). As a result, a switching of the specificity of lysyl-tRNA synthetase from tRNALys (anticodon UUU) toward tRNAAsp (GUC) could be attempted by transplanting the small loop L45 of aspartyl-tRNA synthetase inside lysyl-tRNA synthetase. Upon this transplantation, lysyl-tRNA synthetase loses its capacity to aminoacylate tRNALys. In exchange, the chimeric enzyme acquires the capacity to charge tRNAAsp with lysine. Upon giving the tRNAAsp substrate the discriminator base of tRNALys, the specificity shift is improved. The change of specificity was also established in vivo. Indeed, the transplanted lysyl-tRNA synthetase succeeds in suppressing a missense Lys --> Asp mutation inserted into the beta-lactamase gene. These results functionally establish that sequence variation in a small peptide region of subclass IIb aminoacyl-tRNA synthetases contributes to specification of nucleic acid recognition. Because this peptide element is not part of the core catalytic structure, it may have evolved independently of the active sites of these synthetases. PMID:12766171

  18. Modal wavefront reconstruction over general shaped aperture by numerical orthogonal polynomials

    NASA Astrophysics Data System (ADS)

    Ye, Jingfei; Li, Xinhua; Gao, Zhishan; Wang, Shuai; Sun, Wenqing; Wang, Wei; Yuan, Qun

    2015-03-01

    In practical optical measurements, the wavefront data are recorded by pixelated imaging sensors. The closed-form analytical base polynomial will lose its orthogonality in the discrete wavefront database. For a wavefront with an irregularly shaped aperture, the corresponding analytical base polynomials are laboriously derived. The use of numerical orthogonal polynomials for reconstructing a wavefront with a general shaped aperture over the discrete data points is presented. Numerical polynomials are orthogonal over the discrete data points regardless of the boundary shape of the aperture. The performance of numerical orthogonal polynomials is confirmed by theoretical analysis and experiments. The results demonstrate the adaptability, validity, and accuracy of numerical orthogonal polynomials for estimating the wavefront over a general shaped aperture from regular boundary to an irregular boundary.

  19. Design of generalised orthogonal filters: application to the modelling of dynamical systems

    NASA Astrophysics Data System (ADS)

    Nikolić, Saša S.; Antić, Dragan S.; Perić, Staniša Lj.; Danković, Nikola B.; Milojković, Marko T.

    2016-02-01

    In this article, we define a new class of orthogonal filters with complex poles and zeroes inside their transfer function. This further improvement of classical orthogonal filters allows the possibility to model a wider range of real systems, that is, the systems whose mathematical models have complex zeroes besides real ones. These filters can be applied in the following areas: circuit theory, telecommunications, signal processing, bond graphs, theory approximations and control system theory. First, we describe the rational functions with complex poles and zeroes, and prove their orthogonality. Based on these functions, we designed the block diagram of orthogonal Legendre-type filter with complex poles and zeroes. After that an appropriate analogue scheme of this filter for practical realisation is derived. To validate theoretical results, we performed an experiment with a cascade-connected system designed and practically realised in our laboratories. The experiments proved the quality of the designed orthogonal model in terms of accuracy and simplicity.

  20. S-R compatibility effects with orthogonal stimulus and response dimensions

    NASA Technical Reports Server (NTRS)

    Andre, Anthony D.; Haskell, Ian; Wickens, Christopher D.

    1991-01-01

    An experiment was conducted to assess the relative influence of several factors on performance with orthogonal stimulus-response arrays. Subjects responded to the onset of one of three aligned light circles with a press of one of three aligned response keys. The response array was aligned parallel, angled, or orthogonal to the stimulus array. The results indicated that performance with orthogonal arrays is worse than with parallel or angled S-R arrays. For the orthogonal arrangements, the results also indicate that each hand prefers a mapping directly opposite to the other hand, and that this mapping reverses when the orientation of the stimulus and response arrays are transposed. In addition, the results also revealed that the relative costs of orthogonal S-R arrangements are somewhat attenuated when the assigned mapping associates (i.e., colocates) a given display with its closest control.

  1. 622-Mbps Orthogonal Frequency Division Multiplexing (OFDM) Digital Modem Implemented

    NASA Technical Reports Server (NTRS)

    Kifle, Muli; Bizon, Thomas P.; Nguyen, Nam T.; Tran, Quang K.; Mortensen, Dale J.

    2002-01-01

    Future generation space communications systems feature significantly higher data rates and relatively smaller frequency spectrum allocations than systems currently deployed. This requires the application of bandwidth- and power-efficient signal transmission techniques. There are a number of approaches to implementing such techniques, including analog, digital, mixed-signal, single-channel, or multichannel systems. In general, the digital implementations offer more advantages; however, a fully digital implementation is very difficult because of the very high clock speeds required. Multichannel techniques are used to reduce the sampling rate. One such technique, multicarrier modulation, divides the data into a number of low-rate channels that are stacked in frequency. Orthogonal frequency division multiplexing (OFDM), a form of multicarrier modulation, is being proposed for numerous systems, including mobile wireless and digital subscriber link communication systems. In response to this challenge, NASA Glenn Research Center's Communication Technology Division has developed an OFDM digital modem (modulator and demodulator) with an aggregate information throughput of 622 Mbps. The basic OFDM waveform is constructed by dividing an incoming data stream into four channels, each using either 16- ary quadrature amplitude modulation (16-QAM) or 8-phase shift keying (8-PSK). An efficient implementation for an OFDM architecture is being achieved using the combination of a discrete Fourier transform (DFT) at the transmitter to digitally stack the individual carriers, inverse DFT at the receiver to perform the frequency translations, and a polyphase filter to facilitate the pulse shaping.

  2. Magnetocardiogram measured by fundamental mode orthogonal fluxgate array

    NASA Astrophysics Data System (ADS)

    Karo, Hikaru; Sasada, Ichiro

    2015-05-01

    Magnetocardiography (MCG) of healthy volunteers has been measured by using a fundamental mode orthogonal fluxgate magnetometer array of 32 channels in a magnetic shielded room (MSR). Sensor heads, which are employed, consist of a 45 mm long U-shaped amorphous wire core and a 1000-turn solenoid pick-up coil of 30 mm in length and 3 mm in outer diameter. The excitation current of 100 kHz with large dc bias current is fed directly into wire cores, which are connected in series, whereas the signal detection circuit is provided to each of the sensor heads. A special technique to avoid mutual interaction between sensor heads is implemented, where all the sensor heads are excited synchronously by using a single ac source. A 2-D array having 32 sensors with 4 cm grid spacing was used to measure MCG signals inside an MSR. Measured data from each channel were first filtered (0.16-100 Hz pass band), then averaged for 2 min synchronously with electrocardiogram's peaks taken from both hands. Noise remaining after the average is about 1.8 pTrms for the band-width of 0.16-100 Hz. The QRS complex and the T-wave are clearly detected.

  3. Large Covariance Estimation by Thresholding Principal Orthogonal Complements

    PubMed Central

    Fan, Jianqing; Liao, Yuan; Mincheva, Martina

    2012-01-01

    This paper deals with the estimation of a high-dimensional covariance with a conditional sparsity structure and fast-diverging eigenvalues. By assuming sparse error covariance matrix in an approximate factor model, we allow for the presence of some cross-sectional correlation even after taking out common but unobservable factors. We introduce the Principal Orthogonal complEment Thresholding (POET) method to explore such an approximate factor structure with sparsity. The POET estimator includes the sample covariance matrix, the factor-based covariance matrix (Fan, Fan, and Lv, 2008), the thresholding estimator (Bickel and Levina, 2008) and the adaptive thresholding estimator (Cai and Liu, 2011) as specific examples. We provide mathematical insights when the factor analysis is approximately the same as the principal component analysis for high-dimensional data. The rates of convergence of the sparse residual covariance matrix and the conditional sparse covariance matrix are studied under various norms. It is shown that the impact of estimating the unknown factors vanishes as the dimensionality increases. The uniform rates of convergence for the unobserved factors and their factor loadings are derived. The asymptotic results are also verified by extensive simulation studies. Finally, a real data application on portfolio allocation is presented. PMID:24348088

  4. Aeroelastic System Development Using Proper Orthogonal Decomposition and Volterra Theory

    NASA Technical Reports Server (NTRS)

    Lucia, David J.; Beran, Philip S.; Silva, Walter A.

    2003-01-01

    This research combines Volterra theory and proper orthogonal decomposition (POD) into a hybrid methodology for reduced-order modeling of aeroelastic systems. The out-come of the method is a set of linear ordinary differential equations (ODEs) describing the modal amplitudes associated with both the structural modes and the POD basis functions for the uid. For this research, the structural modes are sine waves of varying frequency, and the Volterra-POD approach is applied to the fluid dynamics equations. The structural modes are treated as forcing terms which are impulsed as part of the uid model realization. Using this approach, structural and uid operators are coupled into a single aeroelastic operator. This coupling converts a free boundary uid problem into an initial value problem, while preserving the parameter (or parameters) of interest for sensitivity analysis. The approach is applied to an elastic panel in supersonic cross ow. The hybrid Volterra-POD approach provides a low-order uid model in state-space form. The linear uid model is tightly coupled with a nonlinear panel model using an implicit integration scheme. The resulting aeroelastic model provides correct limit-cycle oscillation prediction over a wide range of panel dynamic pressure values. Time integration of the reduced-order aeroelastic model is four orders of magnitude faster than the high-order solution procedure developed for this research using traditional uid and structural solvers.

  5. Simultaneous SHG of orthogonally polarized fundamentals in single QPM crystals

    NASA Astrophysics Data System (ADS)

    Johnston, Benjamin F.; Saltiel, Solomon M.; Withford, Michael J.; Kivshar, Yuri S.

    2007-02-01

    Fabrication of quasi-phase-matching (QPM) gratings suitable for cascading of two second-order parametric nonlinear processes in a single lithium niobate crystal is being undertaken using a new technique - electric field poling assisted by laser micro-machined topographical electrodes. To date, single period poled gratings with 45.75, and 45.8 μm periods have been fabricated in order to demonstrate second harmonic generation of 1064nm laser light with 1 st order type-I and 7 th order type-0 QPM simultaneously. The two frequency doubling processes share a common Z polarized second-harmonic wave which allows exchange of energy between the two orthogonally polarized fundamental waves and several second order cascading interactions can be realized. The use of the higher QPM orders (3rd, 5th or 7th) for the type-0 second harmonic generation process leads to comparable efficiencies of the two processes, as the respective nonlinear coefficients are d zzz ~27 pm/V and d yyz ~ 4.7 pm/V in lithium niobate crystals. Possible applications include; polarization switching, parametric amplification and polarization mode dispersion monitoring, and polarization insensitive second harmonic generation.

  6. An index of signal mode complexity based on orthogonal transformation.

    PubMed

    Bhattacharya, Joydeep; Pereda, Ernesto

    2010-08-01

    Irregular and complex signals are ubiquitous in nature. The principal aim of this paper is to develop an index, quantifying the complexity of such signals, which is based on the distribution of the strengths of its orthogonal oscillatory modes estimated by singular value decomposition. The index is first tested with simulated chaotic and/or stochastic maps and flows. Among neural data analysis, the index is first applied to a cognitive EEG data set recorded from two groups, musicians and non-musicians, during listening to music and resting state. In the gamma band (30-50 Hz), musicians showed robust changes in complexity, consistent over various scalp regions, during listening to music from resting condition, whereas such changes were minimal for non-musicians. Then the index is used to separate healthy participants from epileptic and manic patients based on spontaneous EEG analysis. Finally, it is used to track a tonic-clonic seizure EEG signal, and a conspicuous change was found in the complexity profiles of delta band (1-3.5 Hz) oscillations at the onset of seizure. We conclude that this index would be useful for quantification of a wide range of time series including neural oscillations. PMID:19418211

  7. Orthogonal-rotating tetrahedral scanning for cone-beam CT

    NASA Astrophysics Data System (ADS)

    Ye, Ivan B.; Wang, Ge

    2012-10-01

    In this article, a cone-beam CT scanning mode is designed assuming four x-ray sources and a spherical sample. The x-ray sources are mounted at the vertices of a regular tetrahedron. On the circumsphere of the tetrahedron, four detection panels are mounted opposite to each vertex. To avoid x-ray interference, the largest half angle of each x-ray cone beam is 27°22', while the radius of the largest ball fully covered by all the cone beams is 0.460, when the radius of the circumsphere is 1. Several scanning schemes are proposed which consist of two rotations about orthogonal axes, such that each quarter turn provides sufficient data for theoretically exact and stable reconstruction. This design can be used in biomedical or industrial settings, such as when a sequence of reconstructions of an object is desired. Similar scanning schemes based on other regular or irregular polyhedra and various rotation speeds are also discussed.

  8. Dual, orthogonal, backlit pinhole radiography in OMEGA experiments

    NASA Astrophysics Data System (ADS)

    Kuranz, C. C.; Blue, B. E.; Drake, R. P.; Robey, H. F.; Hansen, J. F.; Knauer, J. P.; Grosskopf, M. J.; Krauland, C.; Marion, D. C.

    2006-10-01

    Backlit pinhole radiography used with ungated film as a detector creates x-ray radiographs with increased resolution and contrast. Current hydrodynamics experiments on the OMEGA Laser use a three-dimensional sinusoidal pattern as a seed perturbation for the study of instabilities. The structure of this perturbation makes it highly desirable to obtain two simultaneous orthogonal backlighting views. We accomplished this using two backlit pinholes each mounted 12mm from the target. The pinholes, of varying size and shape, were centered on 5mm square foils of 50μm thick Ta. The backlighting is by K-alpha emission from a 500μm square Ti or Sc foil mounted 500μm from the Ta on a plastic substrate. Four laser beams overfill the metal foil, so that the expanding plastic provides radial tamping of the expanding metal plasma. The resulting x-rays pass through the target onto (ungated) direct exposure film (DEF). Interference between the two views is reduced by using a nose cone in front of the DEF, typically with a 9mm Ta aperture and with magnets to deflect electrons. Comparison of varying types of pinholes and film exposures will be presented from recent experiments as well as an analysis of the background noise created using this experimental technique.

  9. Optimal detection using cyclostationary EOFs[Empirical orthogonal function

    SciTech Connect

    Kim, K.Y.; Wu, Q.

    2000-03-01

    The problem of detecting a climate change signal in the climatological record is of obvious importance in any strategies to understand global climate changes. Atmospheric scientists have applied various statistical techniques to the problem of detecting global warming trend due to increased greenhouse gases. Many climatic and geophysical processes are cyclostationary and exhibit appreciable cyclic (monthly, daily, etc.) variation of their statistics in addition to interannual fluctuations. Utilization of this nested variation of statistics will lead to a better chance of detecting a signal in such a varying background noise field, especially in terms of cyclostationary empirical orthogonal functions, which take the nested periodicity of noise statistics into account. To investigate the improved performance of the cyclostationary approach the developed algorithm is applied to three specific detection examples: El Nino, greenhouse warming, and sunspot fluctuations. In all the test cases, signal-to-noise ratio is raised between 2% and 43% compared with that of a stationary detection technique. The variation of signal strength when a detection filter is constructed based on a different section of modeled noise is within the range of mean signal-to-noise ratio for small to moderate signals. There is a significant variation, however, of signal strength when a detection filter is constructed based on a different model dataset. This implies that model discrepancy is a more important factor than sampling error for the accuracy of the detection method and that climate models need to be improved further in their noise statistics.

  10. Origin of Orthogonality of Strain-Promoted Click Reactions

    PubMed Central

    Wagner, Johannes A; Mercadante, Davide; Nikić, Ivana; Lemke, Edward A; Gräter, Frauke

    2015-01-01

    Site-specific labeling of biomolecules is rapidly advancing due to the discovery of novel mutually orthogonal reactions. Quantum chemistry studies have also increased our understanding of their relative rates, although these have until now been based on highly simplified reactants. Here we examine a set of strain-promoted click-type cycloaddition reactions of n-propyl azide, 3-benzyl tetrazine and 3-benzyl-6-methyl tetrazine with cyclooctenes/ynes, in which we aim to address all relevant structural details of the reactants. Our calculations have included the obligatory handles used to attach the label and biomolecule as these can critically influence the stereochemistry and electron demand of the reaction. We systematically computed orbital gaps, activation and distortion energies using density functional theory and determined experimental rates for validation. Our results challenge the current paradigm of the inverse electron demand for this class of reactions. We found that the ubiquitous handles, when next to the triple bond of cyclooctynes, can switch the Diels–Alder type ligations to normal electron demand, a class we term as SPINEDAC reactions. Electron donating substituents on tetrazine can enhance normal demand but also increase distortion penalties. The presence and isomeric configuration of handles thus determine the reaction speed and regioselectivity. Our findings can be directly utilized in engineering genuine cycloaddition click chemistries for biological labeling. PMID:26178299

  11. Origin of Orthogonality of Strain-Promoted Click Reactions.

    PubMed

    Wagner, Johannes A; Mercadante, Davide; Nikić, Ivana; Lemke, Edward A; Gräter, Frauke

    2015-08-24

    Site-specific labeling of biomolecules is rapidly advancing due to the discovery of novel mutually orthogonal reactions. Quantum chemistry studies have also increased our understanding of their relative rates, although these have until now been based on highly simplified reactants. Here we examine a set of strain-promoted click-type cycloaddition reactions of n-propyl azide, 3-benzyl tetrazine and 3-benzyl-6-methyl tetrazine with cyclooctenes/ynes, in which we aim to address all relevant structural details of the reactants. Our calculations have included the obligatory handles used to attach the label and biomolecule as these can critically influence the stereochemistry and electron demand of the reaction. We systematically computed orbital gaps, activation and distortion energies using density functional theory and determined experimental rates for validation. Our results challenge the current paradigm of the inverse electron demand for this class of reactions. We found that the ubiquitous handles, when next to the triple bond of cyclooctynes, can switch the Diels-Alder type ligations to normal electron demand, a class we term as SPINEDAC reactions. Electron donating substituents on tetrazine can enhance normal demand but also increase distortion penalties. The presence and isomeric configuration of handles thus determine the reaction speed and regioselectivity. Our findings can be directly utilized in engineering genuine cycloaddition click chemistries for biological labeling. PMID:26178299

  12. A Fast Iterated Orthogonal Projection Framework for Smoke Simulation.

    PubMed

    Yang, Yang; Yang, Xubo; Yang, Shuangcai

    2016-05-01

    We present a fast iterated orthogonal projection (IOP) framework for smoke simulations. By modifying the IOP framework with a different means for convergence, our framework significantly reduces the number of iterations required to converge to the desired precision. Our new iteration framework adds a divergence redistributor component to IOP that can improve the impeded convergence logic of IOP. We tested Jacobi, GS and SOR as divergence redistributors and used the Multigrid scheme to generate a highly efficient Poisson solver. It provides a rapid convergence rate and requires less computation time. In all of our experiments, our method only requires 2-3 iterations to satisfy the convergence condition of 1e-5 and 5-7 iterations for 1e-10. Compared with the commonly used Incomplete Cholesky Preconditioned Conjugate Gradient(ICPCG) solver, our Poisson solver accelerates the overall speed to approximately 7- to 30-fold faster for grids ranging from 128(3) to 256(3). Our solver can accelerate more on larger grids because of the property that the iteration count required to satisfy the convergence condition is independent of the problem size. We use various experimental scenes and settings to demonstrate the efficiency of our method. In addition, we present a feasible method for both IOP and our fast IOP to support free surfaces. PMID:27045907

  13. Proper Orthogonal Decomposition in Optimal Control of Fluids

    NASA Technical Reports Server (NTRS)

    Ravindran, S. S.

    1999-01-01

    In this article, we present a reduced order modeling approach suitable for active control of fluid dynamical systems based on proper orthogonal decomposition (POD). The rationale behind the reduced order modeling is that numerical simulation of Navier-Stokes equations is still too costly for the purpose of optimization and control of unsteady flows. We examine the possibility of obtaining reduced order models that reduce computational complexity associated with the Navier-Stokes equations while capturing the essential dynamics by using the POD. The POD allows extraction of certain optimal set of basis functions, perhaps few, from a computational or experimental data-base through an eigenvalue analysis. The solution is then obtained as a linear combination of these optimal set of basis functions by means of Galerkin projection. This makes it attractive for optimal control and estimation of systems governed by partial differential equations. We here use it in active control of fluid flows governed by the Navier-Stokes equations. We show that the resulting reduced order model can be very efficient for the computations of optimization and control problems in unsteady flows. Finally, implementational issues and numerical experiments are presented for simulations and optimal control of fluid flow through channels.

  14. Dual, orthogonal, backlit pinhole radiography in OMEGA experiments

    SciTech Connect

    Kuranz, C. C.; Blue, B. E.; Drake, R. P.; Robey, H. F.; Hansen, J. F.; Knauer, J. P.; Grosskopf, M. J.; Krauland, C.; Marion, D. C.

    2006-10-15

    Backlit pinhole radiography used with ungated film as a detector creates x-ray radiographs with increased resolution and contrast. Current hydrodynamics experiments on the OMEGA Laser use a three-dimensional sinusoidal pattern as a seed perturbation for the study of instabilities. The structure of this perturbation makes it highly desirable to obtain two simultaneous orthogonal backlighting views. We accomplished this using two backlit pinholes each mounted 12 mm from the target. The pinholes, of varying size and shape, were centered on 5 mm square foils of 50 {mu}m thick Ta. The backlighting is by K-alpha emission from a 500 {mu}m square Ti or Sc foil mounted 500 {mu}m from the Ta on a plastic substrate. Four laser beams overfill the metal foil, so that the expanding plastic provides radial tamping of the expanding metal plasma. The resulting x-rays pass through the target onto (ungated) direct exposure film (DEF). Interference between the two views is reduced by using a nose cone in front of the DEF, typically with a 9 mm Ta aperture and with magnets to deflect electrons. Comparison of varying types of pinholes and film exposures will be presented from recent experiments as well as an analysis of the background noise created using this experimental technique.

  15. 622-Mbps Orthogonal Frequency Division Multiplexing Modulator Developed

    NASA Technical Reports Server (NTRS)

    Nguyen, Na T.

    1999-01-01

    The Communications Technology Division at the NASA Lewis Research Center is developing advanced electronic technologies for the space communications and remote sensing systems of tomorrow. As part of the continuing effort to advance the state-of-the art in satellite communications and remote sensing systems, Lewis is developing a programmable Orthogonal Frequency Division Multiplexing (OFDM) modulator card for high-data-rate communication links. The OFDM modulator is particularly suited to high data-rate downlinks to ground terminals or direct data downlinks from near-Earth science platforms. It can support data rates up to 622 megabits per second (Mbps) and high-order modulation schemes such as 16-ary quadrature amplitude modulation (16-ary QAM) or 8- phase shift keying (8PSK). High order modulations can obtain the bandwidth efficiency over the traditional binary phase shift keying (BPSK) or quadrature phase shift keying (QPSK) modulator schemes. The OFDM modulator architecture can also be precompensated for channel disturbances and alleviate amplitude degradations caused by nonlinear transponder characteristics.

  16. [Orthogonal projection divergence-based hyperspectral band selection].

    PubMed

    Su, Hong-jun; Sheng, Ye-hua; Yang, He; Du, Qian

    2011-05-01

    Due to the high data dimensionality of a hyperspectral image, dimensionality reduction algorithm has attracted much attention in hyperspectral image analysis. Band selection algorithm, which selects appropriate bands from the original set of spectral bands, can preserve original information from the data and is useful for image classification and recognition. In the present paper, a novel band selection algorithm based on orthogonal projection divergence (OPD) is proposed, it aims to discriminate the interesting objects from background and noise information, maximize the spectral similarity between different spectral vectors by projecting the original data to feature space. Two HYDICE Washington DC Mall images and an HYMAP Purdue campus image data were experimented, and support vector machine (SVM) classifier was used for classification. The selected band number varies from 5 to 40 in order to study the impacts of different band selection algorithms on different features. For the computation complex, the sequential floating forward search (SFFS) was used to get the appropriate bands. The experiments have proved that our proposed OPD algorithm can outperform other traditional band selection methods such as SAM, ED, SID, and LCMV-BCC for hyperspectral image analysis. It is proven that OPD band selection is effective and robust in hyperspectral remote sensing dimensionality reduction PMID:21800589

  17. Rhythm analysis of orthogonal signals from human walking.

    PubMed

    Ekimov, Alexander; Sabatier, James M

    2011-03-01

    In physical terms, periodic movements of a human body resulting from walking produce a pulse sequence with repetition time T(1) (instant cadence frequency, 1/T(1)) and duration time T(2). Footstep forces generate periodic T(1) broadband seismic and sound signals due to the dynamic forces between the foot and the ground/floor with duration time T(2), which is equal to the time interval for a single footstep from heel strike to toe slap and weight transfer. In a human gait study (for normal speeds of walking), T(1) was detected as 0.5-0.69 s and double limb support takes up about 12% of the gait cycle (2T(1)), so T(2) is greater than 0.12-0.17 s. Short range (of about 50 m) signatures for 30 humans were recorded simultaneously by four orthogonal sensor types at two locations. The sensor types were active Doppler sonar/radar and passive seismic/acoustics. Analysis of signals from these four sensors collected for walking humans showed temporal synchronization and stability of the cadence frequencies, and the cadence frequency from each sensor was equivalent. The time delay between signals from these sensors due to the differences in speeds of propagation for seismic, sound, and electromagnetic waves allows calculation of the distance from a walker to the sensor suite. PMID:21428494

  18. Molecular cloning of the human CTP synthetase gene by functional complementation with purified human metaphase chromosomes.

    PubMed

    Yamauchi, M; Yamauchi, N; Meuth, M

    1990-07-01

    Successive rounds of chromosome-mediated gene transfer were used to complement a hamster cytidine auxotroph deficient in CTP synthetase activity and eventually to clone human genomic and cDNA fragments coding for the structural gene. Our approach was to isolate human Alu+ fragments from a tertiary transfectant and to utilize these fragments to screen a panel of primary transfectants. In this manner two DNA fragments, both mapping within the structural gene, were identified and used to clone a partial length cDNA. The remaining portion of the open reading frame was obtained through the RACE polymerase chain reaction technique. The open reading frame encodes 591 amino acids having a striking degree of similarity to the Escherichia coli structural gene (48% identical amino acids with 76% overall similarity including conservative substitutions) with the glutamine amide transfer domain being particularly conserved. As regulatory mutations of CTP synthetase confer both multi-drug resistance to agents widely used in cancer chemotherapy and a mutator phenotype, the cloning of the structural gene will be important in assessing the relevance of such phenotypes to the development of cellular drug resistance. PMID:2113467

  19. The bifunctional active site of S-adenosylmethionine synthetase. Roles of the basic residues.

    PubMed

    Taylor, J C; Markham, G D

    2000-02-11

    S-adenosylmethionine (AdoMet) synthetase catalyzes a unique two-step enzymatic reaction leading to formation of the primary biological alkylating agent. The crystal structure of Escherichia coli AdoMet synthetase shows that the active site, which lies between two subunits, contains four lysines and one histidine as basic residues. In order to test the proposed charge and hydrogen bonding roles in catalytic function, each lysine has been changed to an uncharged methionine or alanine, and the histidine has been altered to asparagine. The resultant enzyme variants are all tetramers like the wild type enzyme; however, circular dichroism spectra show reductions in helix content for the K245*M and K269M mutants. (The asterisk denotes that the residue is in the second subunit.) Four mutants have k(cat) reductions of approximately 10(3)-10(4)-fold in AdoMet synthesis; however, the k(cat) of K165*M variant is only reduced 2-fold. In each mutant, there is a smaller catalytic impairment in the partial reaction of tripolyphosphate hydrolysis. The K165*A enzyme has a 100-fold greater k(cat) for tripolyphosphate hydrolysis than the wild type enzyme, but this mutant is not activated by AdoMet in contrast to the wild type enzyme. The properties of these mutants require reassessment of the catalytic roles of these residues. PMID:10660564

  20. A human tRNA synthetase is a potent PARP1-activating effector target for resveratrol.

    PubMed

    Sajish, Mathew; Schimmel, Paul

    2015-03-19

    Resveratrol is reported to extend lifespan and provide cardio-neuro-protective, anti-diabetic, and anti-cancer effects by initiating a stress response that induces survival genes. Because human tyrosyl transfer-RNA (tRNA) synthetase (TyrRS) translocates to the nucleus under stress conditions, we considered the possibility that the tyrosine-like phenolic ring of resveratrol might fit into the active site pocket to effect a nuclear role. Here we present a 2.1 Å co-crystal structure of resveratrol bound to the active site of TyrRS. Resveratrol nullifies the catalytic activity and redirects TyrRS to a nuclear function, stimulating NAD(+)-dependent auto-poly-ADP-ribosylation of poly(ADP-ribose) polymerase 1 (PARP1). Downstream activation of key stress signalling pathways are causally connected to TyrRS-PARP1-NAD(+) collaboration. This collaboration is also demonstrated in the mouse, and is specifically blocked in vivo by a resveratrol-displacing tyrosyl adenylate analogue. In contrast to functionally diverse tRNA synthetase catalytic nulls created by alternative splicing events that ablate active sites, here a non-spliced TyrRS catalytic null reveals a new PARP1- and NAD(+)-dependent dimension to the physiological mechanism of resveratrol. PMID:25533949