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Sample records for classical enzyme kinetic

  1. Extending the kinetic solution of the classic Michaelis-Menten model of enzyme action

    OpenAIRE

    BISPO, Jose Ailton Conceicao; Bonafe, Carlos Francisco Sampaio; SOUZA, Volnei Brito de; SILVA, Joao Batista de Almeida e; CARVALHO, Giovani Brandao Mafra de

    2011-01-01

    The principal aim of studies of enzyme-mediated reactions has been to provide comparative and quantitative information on enzyme-catalyzed reactions under distinct conditions. The classic Michaelis-Menten model (Biochem Zeit 49:333, 1913) for enzyme kinetic has been widely used to determine important parameters involved in enzyme catalysis, particularly the Michaelis-Menten constant (K (M) ) and the maximum velocity of reaction (V (max) ). Subsequently, a detailed treatment of the mechanisms ...

  2. Hybrid quantum and classical methods for computing kinetic isotope effects of chemical reactions in solutions and in enzymes.

    Science.gov (United States)

    Gao, Jiali; Major, Dan T; Fan, Yao; Lin, Yen-Lin; Ma, Shuhua; Wong, Kin-Yiu

    2008-01-01

    A method for incorporating quantum mechanics into enzyme kinetics modeling is presented. Three aspects are emphasized: 1) combined quantum mechanical and molecular mechanical methods are used to represent the potential energy surface for modeling bond forming and breaking processes, 2) instantaneous normal mode analyses are used to incorporate quantum vibrational free energies to the classical potential of mean force, and 3) multidimensional tunneling methods are used to estimate quantum effects on the reaction coordinate motion. Centroid path integral simulations are described to make quantum corrections to the classical potential of mean force. In this method, the nuclear quantum vibrational and tunneling contributions are not separable. An integrated centroid path integral-free energy perturbation and umbrella sampling (PI-FEP/UM) method along with a bisection sampling procedure was summarized, which provides an accurate, easily convergent method for computing kinetic isotope effects for chemical reactions in solution and in enzymes. In the ensemble-averaged variational transition state theory with multidimensional tunneling (EA-VTST/MT), these three aspects of quantum mechanical effects can be individually treated, providing useful insights into the mechanism of enzymatic reactions. These methods are illustrated by applications to a model process in the gas phase, the decarboxylation reaction of N-methyl picolinate in water, and the proton abstraction and reprotonation process catalyzed by alanine racemase. These examples show that the incorporation of quantum mechanical effects is essential for enzyme kinetics simulations.

  3. Enzyme kinetics of conjugating enzymes: PAPS sulfotransferase.

    Science.gov (United States)

    James, Margaret O

    2014-01-01

    The sulfotransferase (SULT) enzymes catalyze the formation of sulfate esters or sulfamates from substrates that contain hydroxy or amine groups, utilizing 3'-phosphoadenosyl-5'-phosphosulfate (PAPS) as the donor of the sulfonic group. The rate of product formation depends on the concentrations of PAPS and substrate as well as the sulfotransferase enzyme; thus, if PAPS is held constant while varying substrate concentration (or vice versa), the kinetic constants derived are apparent constants. When studied over a narrow range of substrate concentrations, classic Michaelis-Menten kinetics can be observed with many SULT enzymes and most substrates. Some SULT enzymes exhibit positive or negative cooperativity during conversion of substrate to product, and the kinetics fit the Hill plot. A characteristic feature of most sulfotransferase-catalyzed reactions is that, when studied over a wide range of substrate concentrations, the rate of product formation initially increases as substrate concentration increases, then decreases at high substrate concentrations, i.e., they exhibit substrate inhibition or partial substrate inhibition. This chapter gives an introduction to sulfotransferases, including a historical note, the nomenclature, a description of the function of SULTs with different types of substrates, presentation of examples of enzyme kinetics with SULTs, and a discussion of what is known about mechanisms of substrate inhibition in the sulfotransferases.

  4. Kinetic Measurements for Enzyme Immobilization.

    Science.gov (United States)

    Cooney, Michael J

    2017-01-01

    Enzyme kinetics is the study of the chemical reactions that are catalyzed by enzymes, with a focus on their reaction rates. The study of an enzyme's kinetics considers the various stages of activity, reveals the catalytic mechanism of this enzyme, correlates its value to assay conditions, and describes how a drug or a poison might inhibit the enzyme. Victor Henri initially reported that enzyme reactions were initiated by a bond between the enzyme and the substrate. By 1910, Michaelis and Menten were advancing their work by studying the kinetics of an enzyme saccharase which catalyzes the hydrolysis of sucrose into glucose and fructose. They published their analysis and ever since the Michaelis-Menten equation has been used as the standard to describe the kinetics of many enzymes. Unfortunately, soluble enzymes must generally be immobilized to be reused for long times in industrial reactors. In addition, other critical enzyme properties have to be improved like stability, activity, inhibition by reaction products, and selectivity towards nonnatural substrates. Immobilization is by far the chosen process to achieve these goals.Although the Michaelis-Menten approach has been regularly adapted to the analysis of immobilized enzyme activity, its applicability to the immobilized state is limited by the barriers the immobilization matrix places upon the measurement of compounds that are used to model enzyme kinetics. That being said, the estimated value of the Michaelis-Menten coefficients (e.g., V max, K M) can be used to evaluate effects of immobilization on enzyme activity in the immobilized state when applied in a controlled manner. In this review enzyme activity and kinetics are discussed in the context of the immobilized state, and a few novel protocols are presented that address some of the unique constraints imposed by the immobilization barrier.

  5. The Kinetics of Enzyme Mixtures

    Directory of Open Access Journals (Sweden)

    Simon Brown

    2014-03-01

    Full Text Available Even purified enzyme preparations are often heterogeneous. For example, preparations of aspartate aminotransferase or cytochrome oxidase can consist of several different forms of the enzyme. For this reason we consider how different the kinetics of the reactions catalysed by a mixture of forms of an enzyme must be to provide some indication of the characteristics of the species present. Based on the standard Michaelis-Menten model, we show that if the Michaelis constants (Km of two isoforms differ by a factor of at least 20 the steady-state kinetics can be used to characterise the mixture. However, even if heterogeneity is reflected in the kinetic data, the proportions of the different forms of the enzyme cannot be estimated from the kinetic data alone. Consequently, the heterogeneity of enzyme preparations is rarely reflected in measurements of their steady-state kinetics unless the species present have significantly different kinetic properties. This has two implications: (1 it is difficult, but not impossible, to detect molecular heterogeneity using kinetic data and (2 even when it is possible, a considerable quantity of high quality data is required.

  6. Complex kinetics of fluctuating enzymes: phase diagram characterization of a minimal kinetic scheme.

    Science.gov (United States)

    Min, Wei; Jiang, Liang; Xie, X Sunney

    2010-05-03

    Enzyme molecules are dynamic entities with stochastic fluctuation in both protein conformation and enzymatic activity. However, such a notion of fluctuating enzymes, best characterized by recent single-molecule experiments, was not considered in the classic Michaelis-Menten (MM) kinetic scheme. Here we incorporate the fluctuation concept into the reversible MM scheme, and solve analytically all the possible kinetics (i.e., substrate concentration dependent enzymatic velocity) for a minimal model of fluctuating enzymes. Such a minimal model is found to display a variety of distinct kinetic behaviors (phases) in addition to the classic MM kinetics; excess substrate inhibition, sigmoidal kinetics, and concave biphasic kinetics. We find that all these kinetic phases are interrelated and unified under the framework of fluctuating enzymes and can be adequately described by a phase diagram that consists of two master parameters. Functionally, substrate inhibition, sigmoidal kinetics, and convex biphasic phases exhibit positive cooperativity, whereas concave biphasic phases display negative cooperativity. Remarkably, all these complex kinetics are produced by fluctuating enzymes with single substrate binding site, but the two conformations are, therefore, fundamentally different from the classic MWC and KNF models that require multiple subunit or binding sites. This model also suggests that, for a given enzyme/substrate pair, the non-MM behaviors could undergo transitions among different kinetic phases induced by varying product concentrations, owing to the fundamental Haldane symmetry in the reversible MM scheme.

  7. Extracellular enzyme kinetics scale with resource availability

    Science.gov (United States)

    Microbial community metabolism relies on external digestion, mediated by extracellular enzymes that break down complex organic matter into molecules small enough for cells to assimilate. We analyzed the kinetics of 40 extracellular enzymes that mediate the degradation and assimi...

  8. Enzyme Kinetics and the Michaelis-Menten Equation

    Science.gov (United States)

    Biaglow, Andrew; Erickson, Keith; McMurran, Shawnee

    2010-01-01

    The concepts presented in this article represent the cornerstone of classical mathematical biology. The central problem of the article relates to enzyme kinetics, which is a biochemical system. However, the theoretical underpinnings that lead to the formation of systems of time-dependent ordinary differential equations have been applied widely to…

  9. A Comprehensive Enzyme Kinetic Exercise for Biochemistry

    Science.gov (United States)

    Barton, Janice S.

    2011-01-01

    This article describes a comprehensive treatment of experimental enzyme kinetics strongly coupled to electronic data acquisition and use of spreadsheets to organize data and perform linear and nonlinear least-squares analyses, all in a manner that promotes development of important reasoning skills. Kinetic parameters are obtained for the stable…

  10. Rapid-Equilibrium Enzyme Kinetics

    Science.gov (United States)

    Alberty, Robert A.

    2008-01-01

    Rapid-equilibrium rate equations for enzyme-catalyzed reactions are especially useful because if experimental data can be fit by these simpler rate equations, the Michaelis constants can be interpreted as equilibrium constants. However, for some reactions it is necessary to use the more complicated steady-state rate equations. Thermodynamics is…

  11. Enzyme kinetics: the whole picture reveals hidden meanings.

    Science.gov (United States)

    Pinto, Maria F; Estevinho, Berta N; Crespo, Rosa; Rocha, Fernando A; Damas, Ana M; Martins, Pedro M

    2015-06-01

    The methodology adopted by Michaelis and Menten in 1913 is still routinely used to characterize the catalytic power and selectivity of enzymes. These kinetic measurements must be performed soon after the purified enzyme is mixed with a large excess of substrate. Other time scales and solution compositions are no less physiologically relevant, but fall outside the range of applicability of the classical formalism. Here we show that the complete picture of an enzyme's mode of function is critically obscured by the limited scope of conventional kinetic analysis, even in the simplest case of a single active site without inhibition. This picture is now unveiled in a mathematically closed form that remains valid over the reaction time for all combinations of enzyme/substrate concentrations and rate constants. Algebraic simplicity is maintained in the new formalism when stationary reaction phases are considered. By achieving this century-old objective, the otherwise hidden role of the reversible binding step is revealed and atypical kinetic profiles are explained. Most singular kinetic behaviors are identified in a critical region of conditions that coincide with typical cell conditions. Because it is not covered by the Michaelis-Menten model, the critical region has been missed until now by low- and high-throughput screenings of new drugs. New possibilities are therefore raised for novel and once-promising inhibitors to therapeutically target enzymes.

  12. Enzyme kinetic characterization of protein tyrosine phosphatases

    DEFF Research Database (Denmark)

    Peters, Günther H.J.; Branner, S.; Møller, K. B.

    2003-01-01

    Protein tyrosine phosphatases (PTPs) play a central role in cellular signaling processes, resulting in an increased interest in modulating the activities of PTPs. We therefore decided to undertake a detailed enzyme kinetic evaluation of various transmembrane and cytosolic PTPs (PTPalpha, PTPbeta...

  13. Enzyme kinetics informatics: from instrument to browser.

    Science.gov (United States)

    Swainston, Neil; Golebiewski, Martin; Messiha, Hanan L; Malys, Naglis; Kania, Renate; Kengne, Sylvestre; Krebs, Olga; Mir, Saqib; Sauer-Danzwith, Heidrun; Smallbone, Kieran; Weidemann, Andreas; Wittig, Ulrike; Kell, Douglas B; Mendes, Pedro; Müller, Wolfgang; Paton, Norman W; Rojas, Isabel

    2010-09-01

    A limited number of publicly available resources provide access to enzyme kinetic parameters. These have been compiled through manual data mining of published papers, not from the original, raw experimental data from which the parameters were calculated. This is largely due to the lack of software or standards to support the capture, analysis, storage and dissemination of such experimental data. Introduced here is an integrative system to manage experimental enzyme kinetics data from instrument to browser. The approach is based on two interrelated databases: the existing SABIO-RK database, containing kinetic data and corresponding metadata, and the newly introduced experimental raw data repository, MeMo-RK. Both systems are publicly available by web browser and web service interfaces and are configurable to ensure privacy of unpublished data. Users of this system are provided with the ability to view both kinetic parameters and the experimental raw data from which they are calculated, providing increased confidence in the data. A data analysis and submission tool, the kineticswizard, has been developed to allow the experimentalist to perform data collection, analysis and submission to both data resources. The system is designed to be extensible, allowing integration with other manufacturer instruments covering a range of analytical techniques.

  14. Extracellular enzyme kinetics scale with resource availability

    Science.gov (United States)

    Sinsabaugh, Robert L.; Belnap, Jayne; Findlay, Stuart G.; Follstad Shah, Jennifer J.; Hill, Brian H.; Kuehn, Kevin A.; Kuske, Cheryl; Litvak, Marcy E.; Martinez, Noelle G.; Moorhead, Daryl L.; Warnock, Daniel D.

    2014-01-01

    Microbial community metabolism relies on external digestion, mediated by extracellular enzymes that break down complex organic matter into molecules small enough for cells to assimilate. We analyzed the kinetics of 40 extracellular enzymes that mediate the degradation and assimilation of carbon, nitrogen and phosphorus by diverse aquatic and terrestrial microbial communities (1160 cases). Regression analyses were conducted by habitat (aquatic and terrestrial), enzyme class (hydrolases and oxidoreductases) and assay methodology (low affinity and high affinity substrates) to relate potential reaction rates to substrate availability. Across enzyme classes and habitats, the scaling relationships between apparent Vmax and apparent Km followed similar power laws with exponents of 0.44 to 0.67. These exponents, called elasticities, were not statistically distinct from a central value of 0.50, which occurs when the Km of an enzyme equals substrate concentration, a condition optimal for maintenance of steady state. We also conducted an ecosystem scale analysis of ten extracellular hydrolase activities in relation to soil and sediment organic carbon (2,000–5,000 cases/enzyme) that yielded elasticities near 1.0 (0.9 ± 0.2, n = 36). At the metabolomic scale, the elasticity of extracellular enzymatic reactions is the proportionality constant that connects the C:N:P stoichiometries of organic matter and ecoenzymatic activities. At the ecosystem scale, the elasticity of extracellular enzymatic reactions shows that organic matter ultimately limits effective enzyme binding sites. Our findings suggest that one mechanism by which microbial communities maintain homeostasis is regulating extracellular enzyme expression to optimize the short-term responsiveness of substrate acquisition. The analyses also show that, like elemental stoichiometry, the fundamental attributes of enzymatic reactions can be extrapolated from biochemical to community and ecosystem scales.

  15. Continuum kinetic and multi-fluid simulations of classical sheaths

    Science.gov (United States)

    Cagas, P.; Hakim, A.; Juno, J.; Srinivasan, B.

    2017-02-01

    The kinetic study of plasma sheaths is critical, among other things, to understand the deposition of heat on walls, the effect of sputtering, and contamination of the plasma with detrimental impurities. The plasma sheath also provides a boundary condition and can often have a significant global impact on the bulk plasma. In this paper, kinetic studies of classical sheaths are performed with the continuum kinetic code, Gkeyll, which directly solves the Vlasov-Maxwell equations. The code uses a novel version of the finite-element discontinuous Galerkin scheme that conserves energy in the continuous-time limit. The fields are computed using Maxwell equations. Ionization and scattering collisions are included; however, surface effects are neglected. The aim of this work is to introduce the continuum kinetic method and compare its results with those obtained from an already established finite-volume multi-fluid model also implemented in Gkeyll. Novel boundary conditions on the fluids allow the sheath to form without specifying wall fluxes, so the fluids and fields adjust self-consistently at the wall. The work presented here demonstrates that the kinetic and fluid results are in agreement for the momentum flux, showing that in certain regimes, a multi-fluid model can be a useful approximation for simulating the plasma boundary. There are differences in the electrostatic potential between the fluid and kinetic results. Further, the direct solutions of the distribution function presented here highlight the non-Maxwellian distribution of electrons in the sheath, emphasizing the need for a kinetic model. The densities, velocities, and the potential show a good agreement between the kinetic and fluid results. However, kinetic physics is highlighted through higher moments such as parallel and perpendicular temperatures which provide significant differences from the fluid results in which the temperature is assumed to be isotropic. Besides decompression cooling, the heat flux

  16. Continuum Kinetic and Multi-Fluid Simulations of Classical Sheaths

    CERN Document Server

    Cagas, Petr; Juno, James; Srinivasan, Bhuvana

    2016-01-01

    The kinetic study of plasma sheaths is critical, among other things, to understand the deposition of heat on walls, the effect of sputtering, and contamination of the plasma with detrimental impurities. The plasma sheath also provides a boundary condition and can often have a significant global impact on the bulk plasma. In this paper, kinetic studies of classical sheaths are performed with the continuum code, Gkeyll, that directly solves the Vlasov-Poisson/Maxwell equations. The code uses a novel version of the finite-element discontinuous Galerkin (DG) scheme that conserves energy in the continuous-time limit. The electrostatic field is computed using the Poisson equation. Ionization and scattering collisions are included, however, surface effects are neglected. The aim of this work is to introduce the continuum-kinetic method and compare its results to those obtained from an already established finite-volume multi-fluid model also implemented in Gkeyll. Novel boundary conditions on the fluids allow the she...

  17. Surface enzyme kinetics for biopolymer microarrays: a combination of Langmuir and Michaelis-Menten concepts.

    Science.gov (United States)

    Lee, Hye Jin; Wark, Alastair W; Goodrich, Terry T; Fang, Shiping; Corn, Robert M

    2005-04-26

    Real-time surface plasmon resonance (SPR) imaging measurements of surface enzymatic reactions on DNA microarrays are analyzed using a kinetics model that couples the contributions of both enzyme adsorption and surface enzyme reaction kinetics. For the case of a 1:1 binding of an enzyme molecule (E) to a surface-immobilized substrate (S), the overall enzymatic reaction can be described in terms of classical Langmuir adsorption and Michaelis-Menten concepts and three rate constants: enzyme adsorption (k(a)), enzyme desorption (k(d)) and enzyme catalysis (k(cat)). In contrast to solution enzyme kinetics, the amount of enzyme in solution is in excess as compared to the amount of substrate on the surface. Moreover, the surface concentration of the intermediary enzyme-substrate complex (ES) is not constant with time, but goes to zero as the reaction is completed. However, kinetic simulations show that the fractional surface coverage of ES on the remaining unreacted sites does reach a steady-state value throughout the course of the surface reaction. This steady-state value approaches the Langmuir equilibrium value for cases where k(a)[E] > k(cat). Experiments using the 3' --> 5' exodeoxyribonuclease activity of Exonuclease III on double-stranded DNA microarrays as a function of temperature and enzyme concentration are used to demonstrate how this model can be applied to quantitatively analyze the SPR imaging data.

  18. A century of enzyme kinetic analysis, 1913 to 2013.

    Science.gov (United States)

    Johnson, Kenneth A

    2013-09-02

    This review traces the history and logical progression of methods for quantitative analysis of enzyme kinetics from the 1913 Michaelis and Menten paper to the application of modern computational methods today. Following a brief review of methods for fitting steady state kinetic data, modern methods are highlighted for fitting full progress curve kinetics based upon numerical integration of rate equations, including a re-analysis of the original Michaelis-Menten full time course kinetic data. Finally, several illustrations of modern transient state kinetic methods of analysis are shown which enable the elucidation of reactions occurring at the active sites of enzymes in order to relate structure and function.

  19. A Century of Enzyme Kinetic Analysis, 1913 to 2013

    OpenAIRE

    Johnson, Kenneth A.

    2013-01-01

    This review traces the history and logical progression of methods for quantitative analysis of enzyme kinetics from the 1913 Michaelis and Menten paper to the application of modern computational methods today. Following a brief review of methods for fitting steady state kinetic data, modern methods are highlighted for fitting full progress curve kinetics based upon numerical integration of rate equations, including a re-analysis of the original Michaelis-Menten full time course kinetic data. ...

  20. Enzyme kinetics in drug metabolism: fundamentals and applications.

    Science.gov (United States)

    Nagar, Swati; Argikar, Upendra A; Tweedie, Donald J

    2014-01-01

    Enzymes are protein catalysts that lower the energy barrier for a reaction and speed the rate of a chemical change. The kinetics of reactions catalyzed by enzymes, as well as several mechanisms underlying the kinetics, have been comprehensively studied and written in textbooks (1, 2). The importance of quantitative evaluation of enzymatic processes has been recognized in many fields of study, including biochemistry, molecular biology, and pharmaceutical sciences to name a few. In pharmaceutical sciences, the applications of enzyme kinetics range from hit finding efforts for new chemical entities on a pharmacological target to concentration effect relationships to large-scale biosynthesis. The study of the science of drug metabolism has two principal concepts-rate and extent. While understanding disposition pathways and identification of metabolites provides an insight into the extent of metabolism, kinetics of depletion of substrates (endogenous or exogenous) and formation of metabolites deals with the rate of metabolism. The current textbook specifically focuses on kinetics of drug-metabolizing enzymes, detailing specific enzyme classes, and discusses kinetics as they apply to drug transporters. This textbook also outlines additional factors that contribute to the kinetics of reactions catalyzed by these proteins such as variability in isoforms (pharmacogenomics) and experimental factors including key concepts such as alterations of substrate concentrations due to binding. Applications of these approaches in predicting kinetic parameters and alternative approaches for enzymes (systems biology) and transporters are also discussed. The final section focuses on real-life examples (case studies) to try and exemplify the applications of enzyme kinetic principles. This chapter provides a brief overview outlining some key concepts within each of the sections and the chapters within this textbook.

  1. On the Reducible Character of Haldane-Radić Enzyme Kinetics to Conventional and Logistic Michaelis-Menten Models

    Directory of Open Access Journals (Sweden)

    Mihai V. Putz

    2011-04-01

    Full Text Available The conceptual and practical issues regarding the reduction of the Haldane-Radić enzymic mechanism, specific for cholinesterase kinetics, to the consecrated or logistically modified Michaelis-Menten kinetics, specific for some mutant enzymes, are here clarified as due to the limited initial substrate concentration, through detailed initial rate and progress curve analysis, even when other classical conditions for such equivalence are not entirely fulfilled.

  2. On the reducible character of Haldane-Radić enzyme kinetics to conventional and logistic Michaelis-Menten models.

    Science.gov (United States)

    Putz, Mihai V

    2011-04-13

    The conceptual and practical issues regarding the reduction of the Haldane-Radić enzymic mechanism, specific for cholinesterase kinetics, to the consecrated or logistically modified Michaelis-Menten kinetics, specific for some mutant enzymes, are here clarified as due to the limited initial substrate concentration, through detailed initial rate and progress curve analysis, even when other classical conditions for such equivalence are not entirely fulfilled.

  3. Current IUBMB recommendations on enzyme nomenclature and kinetics

    Directory of Open Access Journals (Sweden)

    Athel Cornish-Bowden

    2014-05-01

    Full Text Available The International Union of Biochemistry (IUB, now IUBMB prepared recommendations for describing the kinetic behaviour of enzymes in 1981. Despite the more than 30 years that have passed since these have not subsequently been revised, though in various respects they do not adequately cover current needs. The IUBMB is also responsible for recommendations on the naming and classification of enzymes. In contrast to the case of kinetics, these recommendations are kept continuously up to date.

  4. Kinetic Monte Carlo simulation of the classical nucleation process

    Science.gov (United States)

    Filipponi, A.; Giammatteo, P.

    2016-12-01

    We implemented a kinetic Monte Carlo computer simulation of the nucleation process in the framework of the coarse grained scenario of the Classical Nucleation Theory (CNT). The computational approach is efficient for a wide range of temperatures and sample sizes and provides a reliable simulation of the stochastic process. The results for the nucleation rate are in agreement with the CNT predictions based on the stationary solution of the set of differential equations for the continuous variables representing the average population distribution of nuclei size. Time dependent nucleation behavior can also be simulated with results in agreement with previous approaches. The method, here established for the case in which the excess free-energy of a crystalline nucleus is a smooth-function of the size, can be particularly useful when more complex descriptions are required.

  5. Reexamining Michaelis-Menten Enzyme Kinetics for Xanthine Oxidase

    Science.gov (United States)

    Bassingthwaighte, James B.; Chinn, Tamara M.

    2013-01-01

    Abbreviated expressions for enzyme kinetic expressions, such as the Michaelis-Menten (M-M) equations, are based on the premise that enzyme concentrations are low compared with those of the substrate and product. When one does progress experiments, where the solute is consumed during conversion to form a series of products, the idealized conditions…

  6. Bioethanol from lignocellulose - pretreatment, enzyme immobilization and hydrolysis kinetics

    DEFF Research Database (Denmark)

    Tsai, Chien Tai

    , the cost of enzyme is still the bottle neck, re-using the enzyme is apossible way to reduce the input of enzyme in the process. In the point view of engineering, the prediction of enzymatic hydrolysis kinetics under different substrate loading, enzyme combination is usful for process design. Therefore...... following enzymatic hydrolysis of hot water pretreated barley straw by immobilized and free BG. Finally, this is the first time that BG aggregates in a calcium alginate were visualized by confocallaser scanning microscope. The images prove that more BG aggregates were entrapped in the matrix when the enzyme...

  7. New types of experimental data shape the use of enzyme kinetics for dynamic network modeling.

    Science.gov (United States)

    Tummler, Katja; Lubitz, Timo; Schelker, Max; Klipp, Edda

    2014-01-01

    Since the publication of Leonor Michaelis and Maude Menten's paper on the reaction kinetics of the enzyme invertase in 1913, molecular biology has evolved tremendously. New measurement techniques allow in vivo characterization of the whole genome, proteome or transcriptome of cells, whereas the classical enzyme essay only allows determination of the two Michaelis-Menten parameters V and K(m). Nevertheless, Michaelis-Menten kinetics are still commonly used, not only in the in vitro context of enzyme characterization but also as a rate law for enzymatic reactions in larger biochemical reaction networks. In this review, we give an overview of the historical development of kinetic rate laws originating from Michaelis-Menten kinetics over the past 100 years. Furthermore, we briefly summarize the experimental techniques used for the characterization of enzymes, and discuss web resources that systematically store kinetic parameters and related information. Finally, describe the novel opportunities that arise from using these data in dynamic mathematical modeling. In this framework, traditional in vitro approaches may be combined with modern genome-scale measurements to foster thorough understanding of the underlying complex mechanisms.

  8. A Comparison of Kinetic Energy and Momentum in Special Relativity and Classical Mechanics

    Science.gov (United States)

    Riggs, Peter J.

    2016-01-01

    Kinetic energy and momentum are indispensable dynamical quantities in both the special theory of relativity and in classical mechanics. Although momentum and kinetic energy are central to understanding dynamics, the differences between their relativistic and classical notions have not always received adequate treatment in undergraduate teaching.…

  9. Extracting enzyme processivity from kinetic assays

    Energy Technology Data Exchange (ETDEWEB)

    Barel, Itay; Brown, Frank L. H. [Department of Chemistry and Biochemistry, University of California, Santa Barbara, California 93106 (United States); Department of Physics, University of California, Santa Barbara, California 93106 (United States); Reich, Norbert O. [Department of Chemistry and Biochemistry, University of California, Santa Barbara, California 93106 (United States)

    2015-12-14

    A steady-state analysis for the catalytic turnover of molecules containing two substrate sites is presented. A broad class of Markovian dynamic models, motivated by the action of DNA modifying enzymes and the rich variety of translocation mechanisms associated with these systems (e.g., sliding, hopping, intersegmental transfer, etc.), is considered. The modeling suggests an elementary and general method of data analysis, which enables the extraction of the enzyme’s processivity directly and unambiguously from experimental data. This analysis is not limited to the initial velocity regime. The predictions are validated both against detailed numerical models and by revisiting published experimental data for EcoRI endonuclease acting on DNA.

  10. Kinetic and thermodynamic aspects of enzyme control and regulation.

    Science.gov (United States)

    Rohwer, Johann M; Hofmeyr, Jan-Hendrik S

    2010-12-16

    This paper develops concepts for assessing and quantifying the regulation of the rate of an enzyme-catalyzed reaction. We show how generic reversible rate equations can be recast in two ways, one making the distance from equilibrium explicit, thereby allowing the distinction between kinetic and thermodynamic control of reaction rate, as well as near-equilibrium and far-from-equilibrium reactions. Recasting in the second form separates mass action from rate capacity and quantifies the degree to which intrinsic mass action contributes to reaction rate and how regulation of an enzyme-catalyzed reaction either enhances or counteracts this mass-action behavior. The contribution of enzyme binding to regulation is analyzed in detail for a number of enzyme-kinetic rate laws, including cooperative reactions.

  11. Measurement of enzyme kinetics and inhibitor constants using enthalpy arrays.

    Science.gov (United States)

    Recht, Michael I; Torres, Frank E; De Bruyker, Dirk; Bell, Alan G; Klumpp, Martin; Bruce, Richard H

    2009-05-15

    Enthalpy arrays enable label-free, solution-based calorimetric detection of molecular interactions in a 96-detector array format. Compared with conventional calorimetry, enthalpy arrays achieve a significant reduction of sample volume and measurement time through the combination of the small size of the detectors and ability to perform measurements in parallel. The current capabilities of the technology for studying enzyme-catalyzed reactions are demonstrated by determining the kinetic parameters for reactions with three model enzymes. In addition, the technology has been used with two classes of enzymes to determine accurate inhibitor constants for competitive inhibitors from measurements at a single inhibitor concentration.

  12. Robust Nonlinear Regression in Enzyme Kinetic Parameters Estimation

    Directory of Open Access Journals (Sweden)

    Maja Marasović

    2017-01-01

    Full Text Available Accurate estimation of essential enzyme kinetic parameters, such as Km and Vmax, is very important in modern biology. To this date, linearization of kinetic equations is still widely established practice for determining these parameters in chemical and enzyme catalysis. Although simplicity of linear optimization is alluring, these methods have certain pitfalls due to which they more often then not result in misleading estimation of enzyme parameters. In order to obtain more accurate predictions of parameter values, the use of nonlinear least-squares fitting techniques is recommended. However, when there are outliers present in the data, these techniques become unreliable. This paper proposes the use of a robust nonlinear regression estimator based on modified Tukey’s biweight function that can provide more resilient results in the presence of outliers and/or influential observations. Real and synthetic kinetic data have been used to test our approach. Monte Carlo simulations are performed to illustrate the efficacy and the robustness of the biweight estimator in comparison with the standard linearization methods and the ordinary least-squares nonlinear regression. We then apply this method to experimental data for the tyrosinase enzyme (EC 1.14.18.1 extracted from Solanum tuberosum, Agaricus bisporus, and Pleurotus ostreatus. The results on both artificial and experimental data clearly show that the proposed robust estimator can be successfully employed to determine accurate values of Km and Vmax.

  13. Oscillatory enzyme reactions and Michaelis-Menten kinetics.

    Science.gov (United States)

    Goldbeter, Albert

    2013-09-02

    Oscillations occur in a number of enzymatic systems as a result of feedback regulation. How Michaelis-Menten kinetics influences oscillatory behavior in enzyme systems is investigated in models for oscillations in the activity of phosphofructokinase (PFK) in glycolysis and of cyclin-dependent kinases in the cell cycle. The model for the PFK reaction is based on a product-activated allosteric enzyme reaction coupled to enzymatic degradation of the reaction product. The Michaelian nature of the product decay term markedly influences the period, amplitude and waveform of the oscillations. Likewise, a model for oscillations of Cdc2 kinase in embryonic cell cycles based on Michaelis-Menten phosphorylation-dephosphorylation kinetics shows that the occurrence and amplitude of the oscillations strongly depend on the ultrasensitivity of the enzymatic cascade that controls the activity of the cyclin-dependent kinase.

  14. [Enzyme kinetics of ligustilide metabolism in rat liver microsomes].

    Science.gov (United States)

    Qian, Min; Shi, Li-fu; Hu, Jin-hong

    2009-04-01

    To study the enzyme kinetics of ligustilide metabolism and the effects of selective CYP450 inhibitors on the metabolism of ligustilide in liver microsomes of rat, a LC-MS method was established for quantitative analysis of ligustilide in liver microsomes incubation system with nitrendipine as internal standard. The determination m/z for ligustilide was 173, and for nitrendipine, 315. An optimum incubation system was found and various selective CYP inhibitors were used to investigate their inhibitory effects on the metabolism of ligustilide. The results showed that enzyme kinetics of ligustilide could be significantly inhibited by ketoconazole, trimethoprim and a-naphthoflavon but scarcely inhibited by omeprazole, 4-methylpyrazole and quinidine. Therefore, CYP3A4, CYP2C9 and CYP1A2 are the major isoenzyme participated in in vitro metabolism of ligustilide.

  15. Evaluation of true kinetic parameters for reversible immobilized enzyme reactions

    Energy Technology Data Exchange (ETDEWEB)

    Ishikawa, H.; Tanaka, T.; Kurose, K.; Hikita, H.

    1987-06-01

    For a reversible one-substrate reaction system that follows the Haldane reaction mechanism, a new and effective method has been proposed to extract true or intrinsic kinetic parameters of immobilized enzymes from diffusion limited rate data. The method utilizes the effectiveness factors correlated in terms of the general modulus defined by Aris and Bischoff, and a new modulus defined in the present study. It requires a trial-and-error calculation, but only a few data points. Furthermore, it provides a saving of materials such as substrates and enzymes, and takes less time for experiments compared to the initial rate methods. The usefulness of the method is demonstrated by determining the kinetic parameters for membrane bound fumarase which catalyzes the reaction of the conversion of fumarate to L-malate, for which the equilibrium constant is circa 4. (Refs. 20).

  16. Parameter estimation for models of ligninolytic and cellulolytic enzyme kinetics

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Gangsheng [ORNL; Post, Wilfred M [ORNL; Mayes, Melanie [ORNL; Frerichs, Joshua T [ORNL; Jagadamma, Sindhu [ORNL

    2012-01-01

    While soil enzymes have been explicitly included in the soil organic carbon (SOC) decomposition models, there is a serious lack of suitable data for model parameterization. This study provides well-documented enzymatic parameters for application in enzyme-driven SOC decomposition models from a compilation and analysis of published measurements. In particular, we developed appropriate kinetic parameters for five typical ligninolytic and cellulolytic enzymes ( -glucosidase, cellobiohydrolase, endo-glucanase, peroxidase, and phenol oxidase). The kinetic parameters included the maximum specific enzyme activity (Vmax) and half-saturation constant (Km) in the Michaelis-Menten equation. The activation energy (Ea) and the pH optimum and sensitivity (pHopt and pHsen) were also analyzed. pHsen was estimated by fitting an exponential-quadratic function. The Vmax values, often presented in different units under various measurement conditions, were converted into the same units at a reference temperature (20 C) and pHopt. Major conclusions are: (i) Both Vmax and Km were log-normal distributed, with no significant difference in Vmax exhibited between enzymes originating from bacteria or fungi. (ii) No significant difference in Vmax was found between cellulases and ligninases; however, there was significant difference in Km between them. (iii) Ligninases had higher Ea values and lower pHopt than cellulases; average ratio of pHsen to pHopt ranged 0.3 0.4 for the five enzymes, which means that an increase or decrease of 1.1 1.7 pH units from pHopt would reduce Vmax by 50%. (iv) Our analysis indicated that the Vmax values from lab measurements with purified enzymes were 1 2 orders of magnitude higher than those for use in SOC decomposition models under field conditions.

  17. Classical Kinetic Theory of Landau Damping for Self-interacting Scalar Fields in the Broken Phase

    OpenAIRE

    1998-01-01

    The classical kinetic theory of one-component self-interacting scalar fields is formulated in the broken symmetry phase and applied to the phenomenon of Landau damping. The domain of validity of the classical approach is found by comparing with the result of a 1-loop quantum calculation.

  18. A Sensitive and Robust Enzyme Kinetic Experiment Using Microplates and Fluorogenic Ester Substrates

    Science.gov (United States)

    Johnson, R. Jeremy; Hoops, Geoffrey C.; Savas, Christopher J.; Kartje, Zachary; Lavis, Luke D.

    2015-01-01

    Enzyme kinetics measurements are a standard component of undergraduate biochemistry laboratories. The combination of serine hydrolases and fluorogenic enzyme substrates provides a rapid, sensitive, and general method for measuring enzyme kinetics in an undergraduate biochemistry laboratory. In this method, the kinetic activity of multiple protein…

  19. Catalyst dynamics: consequences for classical kinetic descriptions of reactors

    DEFF Research Database (Denmark)

    Johannessen, Tue; Larsen, Jane Hvolbæk; Chorkendorff, Ib

    2001-01-01

    The modelling of catalytic reactions/reactors has undergone great improvements since the introduction of empirical power-law kinetics in chemical reaction engineering and micro-kinetic models based on insight into the nature of elementary steps have appeared for many reactions. However, recent...... of the dynamical behaviour of some catalytic systems and discuss the corresponding Limitations in existing models for catalytic reactions and reactors. Catalytic reactors operated in non-steady-state are becoming more frequent in industry. The additional efforts needed to accurately simulate these types...... of reactors are discussed. Finally, we discuss the role of computational fluid dynamics (CFD) as a tool for detailed simulation of catalytic reactors....

  20. Kinetic aspects of human placental alkaline phosphatase enzyme membrane.

    Science.gov (United States)

    Roig, M G; Serrano, M A; Bello, J F; Cachaza, J M; Kennedy, J F

    1991-01-01

    The crosslinking of alkaline phosphatase of human placenta with human serum albumin has been optimized. During the physico-chemical characterization of this immobilized biocatalyst, special attention was paid to attributes such as the irreversibility of the enzyme support bonding, the stability of the catalytic activity, and the effects of pH and temperature on this activity. Regarding stability, patterns of denaturation are proposed, to account for inactivation curves over time and under storage/operation conditions. These patterns, in some cases, indicate the existence of different populations of immobilized enzyme molecules, with a different degree of sensitivity to denaturation. The activity vs pH profiles are clearly modified by the immobilization process. This is because the pH of the free homogeneous solution, measurable with a pH-meter, differs from the real pH of the immediate microenvironment of the immobilized enzyme molecules due to the effects of proton accumulation in the microenvironment (in the reaction catalysed by alkaline phosphatase, protons are produced), to limitations to the free diffusion of H+ and to the possible partition effects of H+ due to polar interactions with residues or molecules of the enzyme membrane. In the experimental working conditions, the apparent optimum temperatures are centered at 40 degrees C, inactivation (thermal denaturation) occurring above this temperature. In the temperature range 10-40 degrees C, the kinetic control over the overall activity of the immobilized enzyme was observed, causing the Arrhenius profiles to be linear.

  1. Kinetics of cassava starch hydrolysis with Termamyl(R) enzyme.

    Science.gov (United States)

    Paolucci-Jeanjean, D; Belleville, M P; Zakhia, N; Rios, G M

    2000-04-05

    Kinetic properties of Termamyl(R) 120L were investigated with respect to starch hydrolysis in a batch reactor at substrate concentrations of 50-270 g. dm(-3) and enzyme concentrations of 0. 17-1 cm(3). dm(-3) (i.e., cm(3) of Novo Nordisk enzymatic solution per dm(3) of raw cassava starch suspension). A general kinetic expression giving product concentration as a function of time was first developed; an equation relating the reaction rate of each product to the sum of the concentrations of oligosaccharides with a higher degree of polymerisation was then derived. The empirical model satisfactorily fits experimental data for oligosaccharides with a degree of polymerization ranging from 1 to 7.

  2. Quantum Corrections to Classical Kinetics: the Weight of Rotation

    CERN Document Server

    Chafin, Clifford

    2016-01-01

    Hydrodynamics of gases in the classical domain are examined from the perspective that the gas has a well-defined wavefunction description at all times. Specifically, the internal energy and volume exclusion of decorrelated vortex structures are included so that quantum corrections and modifications to Navier-Stokes behavior can be derived. This leads to a small deviation in rigid body rotation for a cylindrically bound gas and the internal energy changes associated with vorticity give deviations in the Reynolds' transport theorem. Some macroscopic observable features arising from this include variations in the specific heat, an anisotropic correction to thermal conductivity and a variation in optical scattering as a function of the declination from the axis of local vorticity. The improvements in magneto-optical traps suggests some interesting experiments to be done in higher temperature regimes where they are not usually employed. It is argued that the finite lifetime of observed vortices in ultracold bosoni...

  3. In vitro-in vivo scaling of CYP kinetic data not consistent with the classical Michaelis-Menten model.

    Science.gov (United States)

    Houston, J B; Kenworthy, K E

    2000-03-01

    Strategies for the prediction of in vivo drug clearance from in vitro drug metabolite kinetic data are well established for the rat. In this animal species, metabolism rate-substrate concentration relationships can commonly be described by the classic hyperbola consistent with the Michaelis-Menten model and simple scaling of the parameter intrinsic clearance (CL(int) - the ratio of V(max) to K(m)) is particularly valuable. The in vitro scaling of kinetic data from human tissue is more complex, particularly as many substrates for cytochrome P450 (CYP) 3A4, the dominant human CYP, show nonhyperbolic metabolism rate-substrate concentration curves. This review critically examines these types of data, which require the adoption of an enzyme model with multiple sites showing cooperative binding for the drug substrate, and considers the constraints this kinetic behavior places on the prediction of in vivo pharmacokinetic characteristics, such as metabolic stability and inhibitory drug interaction potential. The cases of autoactivation and autoinhibition are discussed; the former results in an initial lag in the rate-substrate concentration profile to generate a sigmoidal curve whereas the latter is characterized by a convex curve as V(max) is not maintained at high substrate concentrations. When positive cooperativity occurs, we suggest the use of CL(max), the maximal clearance resulting from autoactivation, as a substitute for CL(int). The impact of heteroactivation on this approach is also of importance. In the case of negative cooperativity, care in using the V(max)/K(m) approach to CL(int) determination must be taken. Examples of substrates displaying each type of kinetic behavior are discussed for various recombinant CYP enzymes, and possible artifactual sources of atypical rate-concentration curves are outlined. Finally, the consequences of ignoring atypical Michaelis-Menten kinetic relationships are examined, and the inconsistencies reported for both different

  4. Classic hydrodynamic and kinetic formalism as averaging of delta-functional particle images

    CERN Document Server

    Kuz'menkov, L S

    2014-01-01

    Critical analyses of well-known methods of derivation of kinetic and hydrodynamic equations is presented. Another method of derivation of kinetic and hydrodynamic equations from classic mechanics is described. It is shown that equations of classic hydrodynamics can be derived directly from microscopic picture of motion, without using of kinetic equations as an intermediate step. New method of derivation of equation of macroscopic motion includes explicit averaging of microscopic motion on infinitesimally small piece of medium. This averaging leads to presence of electric dipole, magnetic dipole, and higher moments along with the charge density and the current density in hydrodynamic equations. The method under consideration allows to derive equations of evolution for new quantities.

  5. Parallel versus Off-Pathway Michaelis-Menten Mechanism for Single-Enzyme Kinetics of a Fluctuating Enzyme.

    Science.gov (United States)

    Kumar, Ashutosh; Maity, Hiranmay; Dua, Arti

    2015-07-09

    Recent fluorescence spectroscopy measurements of the turnover time distribution of single-enzyme turnover kinetics of β-galactosidase provide evidence of Michaelis-Menten kinetics at low substrate concentration. However, at high substrate concentrations, the dimensionless variance of the turnover time distribution shows systematic deviations from the Michaelis-Menten prediction. This difference is attributed to conformational fluctuations in both the enzyme and the enzyme-substrate complex and to the possibility of both parallel- and off-pathway kinetics. Here, we use the chemical master equation to model the kinetics of a single fluctuating enzyme that can yield a product through either parallel- or off-pathway mechanisms. An exact expression is obtained for the turnover time distribution from which the mean turnover time and randomness parameters are calculated. The parallel- and off-pathway mechanisms yield strikingly different dependences of the mean turnover time and the randomness parameter on the substrate concentration. In the parallel mechanism, the distinct contributions of enzyme and enzyme-substrate fluctuations are clearly discerned from the variation of the randomness parameter with substrate concentration. From these general results, we conclude that an off-pathway mechanism, with substantial enzyme-substrate fluctuations, is needed to rationalize the experimental findings of single-enzyme turnover kinetics of β-galactosidase.

  6. Parallel versus off-pathway Michaelis-Menten mechanism for single-enzyme kinetics of a fluctuating enzyme

    CERN Document Server

    Kumar, Ashutosh; Dua, Arti

    2015-01-01

    Recent fluorescence spectroscopy measurements of the turnover time distribution of single-enzyme turnover kinetics of $\\beta$-galactosidase provide evidence of Michaelis-Menten kinetics at low substrate concentration. However, at high substrate concentrations, the dimensionless variance of the turnover time distribution shows systematic deviations from the Michaelis-Menten prediction. This difference is attributed to conformational fluctuations in both the enzyme and the enzyme-substrate complex and to the possibility of both parallel and off-pathway kinetics. Here, we use the chemical master equation to model the kinetics of a single fluctuating enzyme that can yield a product through either parallel or off-pathway mechanisms. An exact expression is obtained for the turnover time distribution from which the mean turnover time and randomness parameters are calculated. The parallel and off-pathway mechanisms yield strikingly different dependences of the mean turnover time and the randomness parameter on the su...

  7. Measurement of Enzyme Kinetics by Use of a Blood Glucometer: Hydrolysis of Sucrose and Lactose

    Science.gov (United States)

    Heinzerling, Peter; Schrader, Frank; Schanze, Sascha

    2012-01-01

    An alternative analytical method for measuring the kinetic parameters of the enzymes invertase and lactase is described. Invertase hydrolyzes sucrose to glucose and fructose and lactase hydrolyzes lactose to glucose and galactose. In most enzyme kinetics studies, photometric methods or test strips are used to quantify the derivates of the…

  8. Cadmium phytotoxicity: Quantitative sensitivity relationships between classical endpoints and antioxidative enzyme biomarkers

    Energy Technology Data Exchange (ETDEWEB)

    Rosa Correa, Albertina Xavier da [Centro de Ciencias Tecnologicas da Terra e do Mar, Universidade do Vale do Itajai, Rua Uruguai, 458, 88302-202 Itajai SC (Brazil); Roerig, Leonardo Rubi [Centro de Ciencias Tecnologicas da Terra e do Mar, Universidade do Vale do Itajai, Rua Uruguai, 458, 88302-202 Itajai SC (Brazil); Verdinelli, Miguel A. [Centro de Ciencias Tecnologicas da Terra e do Mar, Universidade do Vale do Itajai, Rua Uruguai, 458, 88302-202 Itajai SC (Brazil); Cotelle, Sylvie [Centre des Sciences de l' Environnement, Universite de Metz, 57000 Metz (France); Ferard, Jean-Francois [Centre des Sciences de l' Environnement, Universite de Metz, 57000 Metz (France); Radetski, Claudemir Marcos [Centro de Ciencias Tecnologicas da Terra e do Mar, Universidade do Vale do Itajai, Rua Uruguai, 458, 88302-202 Itajai SC (Brazil)]. E-mail: radetski@univali.br

    2006-03-15

    In this work, cadmium phytotoxicity and quantitative sensitivity relationships between different hierarchical endpoints in plants cultivated in a contaminated soil were studied. Thus, germination rate, biomass growth and antioxidative enzyme activity (i.e. superoxide dismutase, peroxidase, catalase and glutathione reductase) in three terrestrial plants (Avena sativa L., Brassica campestris L. cv. Chinensis, Lactuca sativa L. cv. hanson) were analyzed. Plant growth tests were carried out according to an International Standard Organization method and the results were analyzed by ANOVA followed by Williams' test. The concentration of Cd{sup 2+} that had the smallest observed significant negative effect (LOEC) on plant biomass was 6.25, 12.5 and 50 mg Cd/kg dry soil for lettuce, oat and Chinese cabbage, respectively. Activity of all enzymes studied increased significantly compared to enzyme activity in plant controls. For lettuce, LOEC values (mg Cd/kg dry soil) for enzymic activity ranged from 0.05 (glutathione reductase) to 0.39 (catalase). For oat, LOEC values (mg Cd/kg dry soil) ranged from 0.19 (for superoxide dismutase and glutathione reductase) to 0.39 (for catalase and peroxidase). For Chinese cabbage, LOEC values (mg Cd/kg dry soil) ranged from 0.19 (peroxidase, catalase and glutathione reductase) to 0.39 (superoxide dismutase). Classical (i.e. germination and biomass) and biochemical (i.e. enzyme activity) endpoints were compared to establish a sensitivity ranking, which was: enzyme activity > biomass > germination rate. For cadmium-soil contamination, the determination of quantitative sensitivity relationships (QSR) between classical and antioxidative enzyme biomarkers showed that the most sensitive plant species have, generally, the lowest QSR values.

  9. Enzyme kinetics of hevamine, a chitinase from the rubber tree Hevea brasiliensis

    NARCIS (Netherlands)

    Bokma, Evert; Barends, Thomas; Terwisscha van Scheltinga, Anke C.; Dijkstra, Bauke W.; Beintema, Jaap J.

    2000-01-01

    The enzyme kinetics of hevamine, a chitinase from the rubber tree Hevea brasiliensis, were studied in detail with a new enzyme assay. In this assay, the enzyme reaction products were derivatized by reductive coupling to a chromophore, Products mere separated by HPLC and the amount of product was cal

  10. A Hands-On Classroom Simulation to Demonstrate Concepts in Enzyme Kinetics

    Science.gov (United States)

    Junker, Matthew

    2010-01-01

    A classroom exercise is described to introduce enzyme kinetics in an undergraduate biochemistry or chemistry course. The exercise is a simulation in which a student acts as an enzyme that "catalyzes" the unscrewing of a nut from a bolt. With other students assisting, the student enzyme carries out reactions with bolt-nut substrates under different…

  11. The use of PAMAM dendrimers as a platform for laccase immobilization: kinetic characterization of the enzyme.

    Science.gov (United States)

    Cardoso, Franciane Pinheiro; Aquino Neto, Sidney; Ciancaglini, Pietro; de Andrade, Adalgisa R

    2012-08-01

    The kinetic behavior of the enzyme laccase in solution and immobilized onto carbon platforms using poly(amido amine) (PAMAM) dendrimers has been investigated. The results with the immobilized enzymes have demonstrated that almost ten times more enzyme on the carbon support is required for satisfactory kinetic rates to be achieved. Furthermore, the study as a function of the substrate concentration revealed that the kinetic behavior of the enzyme in solution fits the Michaelis-Menten model. However, when the enzyme is immobilized onto the carbon surface, the catalyzed reaction follows a particular kinetic behavior with apparent positive cooperativity. The highest activity with laccase (in solution or immobilized) is achieved around pH 4.5, and the substrate conversion rate clearly diminishes with rising pH. The optimum temperature lies around 60 °C. The enzyme displays good catalytic activity in a wide range of pH and temperature values. The stability tests evidenced that there is no appreciable reduction in the enzymatic activity after immobilization within the first 30 days. Taking into account both the kinetic and stability tests, one can infer that the use of PAMAM dendrimers seems to be a very attractive approach for the immobilization of enzymes, as well as a feasible and useful methodology for the anchoring of enzymes with potential application in many biotechnological areas.

  12. Emergence of dynamic cooperativity in the stochastic kinetics of fluctuating enzymes.

    Science.gov (United States)

    Kumar, Ashutosh; Chatterjee, Sambarta; Nandi, Mintu; Dua, Arti

    2016-08-28

    Dynamic co-operativity in monomeric enzymes is characterized in terms of a non-Michaelis-Menten kinetic behaviour. The latter is believed to be associated with mechanisms that include multiple reaction pathways due to enzymatic conformational fluctuations. Recent advances in single-molecule fluorescence spectroscopy have provided new fundamental insights on the possible mechanisms underlying reactions catalyzed by fluctuating enzymes. Here, we present a bottom-up approach to understand enzyme turnover kinetics at physiologically relevant mesoscopic concentrations informed by mechanisms extracted from single-molecule stochastic trajectories. The stochastic approach, presented here, shows the emergence of dynamic co-operativity in terms of a slowing down of the Michaelis-Menten (MM) kinetics resulting in negative co-operativity. For fewer enzymes, dynamic co-operativity emerges due to the combined effects of enzymatic conformational fluctuations and molecular discreteness. The increase in the number of enzymes, however, suppresses the effect of enzymatic conformational fluctuations such that dynamic co-operativity emerges solely due to the discrete changes in the number of reacting species. These results confirm that the turnover kinetics of fluctuating enzyme based on the parallel-pathway MM mechanism switches over to the single-pathway MM mechanism with the increase in the number of enzymes. For large enzyme numbers, convergence to the exact MM equation occurs in the limit of very high substrate concentration as the stochastic kinetics approaches the deterministic behaviour.

  13. Emergence of dynamic cooperativity in the stochastic kinetics of fluctuating enzymes

    Science.gov (United States)

    Kumar, Ashutosh; Chatterjee, Sambarta; Nandi, Mintu; Dua, Arti

    2016-08-01

    Dynamic co-operativity in monomeric enzymes is characterized in terms of a non-Michaelis-Menten kinetic behaviour. The latter is believed to be associated with mechanisms that include multiple reaction pathways due to enzymatic conformational fluctuations. Recent advances in single-molecule fluorescence spectroscopy have provided new fundamental insights on the possible mechanisms underlying reactions catalyzed by fluctuating enzymes. Here, we present a bottom-up approach to understand enzyme turnover kinetics at physiologically relevant mesoscopic concentrations informed by mechanisms extracted from single-molecule stochastic trajectories. The stochastic approach, presented here, shows the emergence of dynamic co-operativity in terms of a slowing down of the Michaelis-Menten (MM) kinetics resulting in negative co-operativity. For fewer enzymes, dynamic co-operativity emerges due to the combined effects of enzymatic conformational fluctuations and molecular discreteness. The increase in the number of enzymes, however, suppresses the effect of enzymatic conformational fluctuations such that dynamic co-operativity emerges solely due to the discrete changes in the number of reacting species. These results confirm that the turnover kinetics of fluctuating enzyme based on the parallel-pathway MM mechanism switches over to the single-pathway MM mechanism with the increase in the number of enzymes. For large enzyme numbers, convergence to the exact MM equation occurs in the limit of very high substrate concentration as the stochastic kinetics approaches the deterministic behaviour.

  14. Flow-Based Systems for Rapid and High-Precision Enzyme Kinetics Studies

    Directory of Open Access Journals (Sweden)

    Supaporn Kradtap Hartwell

    2012-01-01

    Full Text Available Enzyme kinetics studies normally focus on the initial rate of enzymatic reaction. However, the manual operation of steps of the conventional enzyme kinetics method has some drawbacks. Errors can result from the imprecise time control and time necessary for manual changing the reaction cuvettes into and out of the detector. By using the automatic flow-based analytical systems, enzyme kinetics studies can be carried out at real-time initial rate avoiding the potential errors inherent in manual operation. Flow-based systems have been developed to provide rapid, low-volume, and high-precision analyses that effectively replace the many tedious and high volume requirements of conventional wet chemistry analyses. This article presents various arrangements of flow-based techniques and their potential use in future enzyme kinetics applications.

  15. Emergence of Dynamic Cooperativity in the Stochastic Kinetics of Fluctuating Enzymes

    CERN Document Server

    Kumar, Ashutosh; Nandi, Mintu; Dua, Arti

    2016-01-01

    Dynamic cooperativity in monomeric enzymes is characterized in terms of a non-Michaelis-Menten kinetic behaviour. The latter is believed to be associated with mechanisms that include multiple reaction pathways due to enzymatic conformational fluctuations. Recent advances in single-molecule fluorescence spectroscopy have provided new fundamental insights on the possible mechanisms underlying reactions catalyzed by fluctuating enzymes. Here, we present a bottom-up approach to understand enzyme turnover kinetics at physiologically relevant mesoscopic concentrations informed by mechanisms extracted from single-molecule stochastic trajectories. The stochastic approach, presented here, shows the emergence of dynamic cooperativity in terms of a slowing down of the Michaelis-Menten (MM) kinetics resulting in negative cooperativity. For fewer enzymes, dynamic cooperativity emerges due to the combined effects of enzymatic conformational fluctuations and molecular discreteness. The increase in the number of enzymes, how...

  16. ENZYME KINETICS FOR SYSTEMS BIOLOGY : WHEN, WHY AND HOW

    NARCIS (Netherlands)

    Adamczyk, Malgorzata; van Eunen, Karen; Bakker, Barbara M.; Westerhoff, Hans V.; Jameson, D; Verma, M; Westerhoff, HV

    2011-01-01

    In vitro enzymatic assays of cell-free extracts offer an opportunity to assess in vivo enzyme concentrations. If performed under conditions that resemble the conditions in vivo, they may also reveal some of the capacities and properties of the same enzymes in vivo; we shall call this the ex vivo app

  17. The Nuts and Bolts of Michaelis-Menten Enzyme Kinetics: Suggestions and Clarifications

    Science.gov (United States)

    Silverstein, Todd

    2011-01-01

    Matthew Junker's recent article describes a useful and effective enzyme kinetics application and analogy in which students simulate enzyme activity by unscrewing nut-bolt "substrate molecules", thus, converting them into separate nuts and bolts "products". A number of suggestions and corrections are presented that improve the clarity and accuracy…

  18. Simplified yet highly accurate enzyme kinetics for cases of low substrate concentrations

    NARCIS (Netherlands)

    Härdin, H.M.; Zagaris, A.; Krab, K.; Westerhoff, H.V.

    2009-01-01

    Much of enzyme kinetics builds on simplifications enabled by the quasi-steady-state approximation and is highly useful when the concentration of the enzyme is much lower than that of its substrate. However, in vivo, this condition is often violated. In the present study, we show that, under conditio

  19. Studies on Enzyme Kinetics by Microchip and Related Techniques

    Institute of Scientific and Technical Information of China (English)

    James J. Bao; WANG Huai-Feng; ZHOU Da-Wei; Ken R. Wehmeyer

    2004-01-01

    Both conventional and microchip-based capillary electrophoresis(CE) technologies have been used for the analysis of enzymes. Practical procedures of using CE to determine the Km and Vmax values of an enzyme have been developed. By studying the inhibition to the enzyme, it is possible to select a suitable drug candidate. When compared with the conventional CE method, single lane microchip-based method can improve the speed for the assay three times. By using multiple lane-based microchip, the speed can be further increased.

  20. More Nuts and Bolts of Michaelis-Menten Enzyme Kinetics

    Science.gov (United States)

    Lechner, Joseph H.

    2011-01-01

    Several additions to a classroom activity are proposed in which an "enzyme" (the student) converts "substrates" (nut-bolt assemblies) into "products" (separated nuts and bolts) by unscrewing them. (Contains 1 table.)

  1. An Enzyme Kinetics Experiment for the Undergraduate Organic Chemistry Laboratory

    Science.gov (United States)

    Olsen, Robert J.; Olsen, Julie A.; Giles, Greta A.

    2010-01-01

    An experiment using [superscript 1]H NMR spectroscopy to observe the kinetics of the acylase 1-catalyzed hydrolysis of "N"-acetyl-DL-methionine has been developed for the organic laboratory. The L-enantiomer of the reactant is hydrolyzed completely in less than 2 h, and [superscript 1]H NMR spectroscopic data from a single sample can be worked up…

  2. Enzyme kinetics and the maximum entropy production principle.

    Science.gov (United States)

    Dobovišek, Andrej; Zupanović, Paško; Brumen, Milan; Bonačić-Lošić, Zeljana; Kuić, Domagoj; Juretić, Davor

    2011-03-01

    A general proof is derived that entropy production can be maximized with respect to rate constants in any enzymatic transition. This result is used to test the assumption that biological evolution of enzyme is accompanied with an increase of entropy production in its internal transitions and that such increase can serve to quantify the progress of enzyme evolution. The state of maximum entropy production would correspond to fully evolved enzyme. As an example the internal transition ES↔EP in a generalized reversible Michaelis-Menten three state scheme is analyzed. A good agreement is found among experimentally determined values of the forward rate constant in internal transitions ES→EP for three types of β-Lactamase enzymes and their optimal values predicted by the maximum entropy production principle, which agrees with earlier observations that β-Lactamase enzymes are nearly fully evolved. The optimization of rate constants as the consequence of basic physical principle, which is the subject of this paper, is a completely different concept from a) net metabolic flux maximization or b) entropy production minimization (in the static head state), both also proposed to be tightly connected to biological evolution.

  3. Renormalized kinetic theory of classical fluids in and out of equilibrium

    CERN Document Server

    Daligault, Jerome

    2011-01-01

    We present a theory for the construction of renormalized kinetic equations to describe the dynamics of classical systems of particles in or out of equilibrium. A closed, self-consistent set of evolution equations is derived for the single-particle phase-space distribution function $f$, the correlation function $C=$, the retarded and advanced density response functions $\\chi^{R,A}=\\delta f/\\delta\\phi$ to an external potential $\\phi$, and the associated memory functions $\\Sigma^{R,A,C}$. The basis of the theory is an effective action functional $\\Omega$ of external potentials $\\phi$ that contains all information about the dynamical properties of the system. In particular, its functional derivatives generate successively the single-particle phase-space density $f$ and all the correlation and density response functions, which are coupled through an infinite hierarchy of evolution equations. Traditional renormalization techniques are then used to perform the closure of the hierarchy through memory functions. The l...

  4. Enzyme kinetics determined by single-injection isothermal titration calorimetry.

    Science.gov (United States)

    Transtrum, Mark K; Hansen, Lee D; Quinn, Colette

    2015-04-01

    The purposes of this paper are (a) to examine the effect of calorimeter time constant (τ) on heat rate data from a single enzyme injection into substrate in an isothermal titration calorimeter (ITC), (b) to provide information that can be used to predict the optimum experimental conditions for determining the rate constant (k2), Michaelis constant (KM), and enthalpy change of the reaction (ΔRH), and (c) to describe methods for evaluating these parameters. We find that KM, k2 and ΔRH can be accurately estimated without correcting for the calorimeter time constant, τ, if (k2E/KM), where E is the total active enzyme concentration, is between 0.1/τ and 1/τ and the reaction goes to at least 99% completion. If experimental conditions are outside this domain and no correction is made for τ, errors in the inferred parameters quickly become unreasonable. A method for fitting single-injection data to the Michaelis-Menten or Briggs-Haldane model to simultaneously evaluate KM, k2, ΔRH, and τ is described and validated with experimental data. All four of these parameters can be accurately inferred provided the reaction time constant (k2E/KM) is larger than 1/τ and the data include enzyme saturated conditions.

  5. Classical nucleation theory of immersion freezing: sensitivity of contact angle schemes to thermodynamic and kinetic parameters

    Science.gov (United States)

    Ickes, Luisa; Welti, André; Lohmann, Ulrike

    2017-02-01

    Heterogeneous ice formation by immersion freezing in mixed-phase clouds can be parameterized in general circulation models (GCMs) by classical nucleation theory (CNT). CNT parameterization schemes describe immersion freezing as a stochastic process, including the properties of insoluble aerosol particles in the droplets. There are different ways to parameterize the properties of aerosol particles (i.e., contact angle schemes), which are compiled and tested in this paper. The goal of this study is to find a parameterization scheme for GCMs to describe immersion freezing with the ability to shift and adjust the slope of the freezing curve compared to homogeneous freezing to match experimental data. We showed in a previous publication that the resulting freezing curves from CNT are very sensitive to unconstrained kinetic and thermodynamic parameters in the case of homogeneous freezing. Here we investigate how sensitive the outcome of a parameter estimation for contact angle schemes from experimental data is to unconstrained kinetic and thermodynamic parameters. We demonstrate that the parameters describing the contact angle schemes can mask the uncertainty in thermodynamic and kinetic parameters. Different CNT formulations are fitted to an extensive immersion freezing dataset consisting of size-selected measurements as a function of temperature and time for different mineral dust types, namely kaolinite, illite, montmorillonite, microcline (K-feldspar), and Arizona test dust. We investigated how accurate different CNT formulations (with estimated fit parameters for different contact angle schemes) reproduce the measured freezing data, especially the time and particle size dependence of the freezing process. The results are compared to a simplified deterministic freezing scheme. In this context, we evaluated which CNT-based parameterization scheme able to represent particle properties is the best choice to describe immersion freezing in a GCM.

  6. Universal Bound on the Fano Factor in Enzyme Kinetics

    CERN Document Server

    Barato, Andre C

    2015-01-01

    The Fano factor, an observable quantifying fluctuations of product generation by a single enzyme, can reveal information about the underlying reaction scheme. A lower bound on this Fano factor that depends on the thermodynamic affinity driving the transformation from substrate to product constrains the number of intermediate states of an enzymatic cycle. So far, this bound has been proven only for a unicyclic network of states. We show that the bound can be extended to arbitrary multicyclic networks, with the Fano factor constraining the largest value of the effective length, which is the ratio between the number of states and the number of products, among all cycles.

  7. Kinetics of soil enzyme activities under different ecosystems: An index of soil quality

    Directory of Open Access Journals (Sweden)

    Monty Kujur

    2014-03-01

    Full Text Available Soil microbial activity plays an important role in regulating biotransformation, nutrient cycling and hence the microbiological processes are at the center of many ecological functions. The kinetic parameters (Vmax and KmMichaelis constant of different enzymes (amylase, invertase, protease, urease, and dehydrogenase were determined in order to assess the metabolic response of soil. The maximum reaction velocity (Vmax represents a maximum rate of activity when all enzymes are saturated, which markedly increased in forest soil as compared to fresh mine spoil due to the gradual accumulation of soil organic matter. Smaller Km value was estimated in forest soil (FS as compared to fresh mine spoil (FMS, suggesting the greater affinity of soil enzymes for substrate in FS. The catalytic efficiency (Vmax /Km reflects an impression on microbial community composition with a change in soil enzymes. These enzyme characters (activities and kinetic parameters have greater significance as early and sensitive indicators of the changes in soil properties induced by different management systems. These parameters (Vmax and Km can be useful markers to assess changes in microbial activity of soil, since they represent quantity and affinity of enzymes respectively. The metabolic index (dehydrogenase activity/organic carbon (OC was found to be correlated with Vmax of dehydrogenase (r = 0.953; p < 0.01 and OC (r = 0.880; p < 0.01. Principal component analysis was able to discriminate seven different soil samples into seven independent clusters based on their enzyme activities and kinetic parameters. Indeed, the study revealed the importance of kinetics study of soil enzymes, which can be considered valid parameters to monitor the evolution of microbiological activity in soil, and hence an index of soil quality.

  8. Single-molecule enzymology of steroid transforming enzymes: Transient kinetic studies and what they tell us.

    Science.gov (United States)

    Penning, Trevor M

    2016-07-01

    Structure-function studies on steroid transforming enzymes often use site-directed mutagenesis to inform mechanisms of catalysis and effects on steroid binding, and data are reported in terms of changes in steady state kinetic parameters kcat, Km and kcat/Km. However, this dissection of function is limited since kcat is governed by the rate-determining step and Km is a complex macroscopic kinetic constant. Often site-directed mutagenesis can lead to a change in the rate-determining step which cannot be revealed by just reporting a decrease in kcat alone. These issues are made more complex when it is considered that many steroid transforming enzymes have more than one substrate and product. We present the case for using transient-kinetics performed with stopped-flow spectrometry to assign rate constants to discrete steps in these multi-substrate reactions and their use to interpret enzyme mechanism and the effects of disease and engineered mutations. We demonstrate that fluorescence kinetic transients can be used to measure ligand binding that may be accompanied by isomerization steps, revealing the existence of new enzyme intermediates. We also demonstrate that single-turnover reactions can provide a klim for the chemical step and Ks for steroid-substrate binding and that when coupled with kinetic isotope effect measurements can provide information on transition state intermediates. We also demonstrate how multiple turnover experiments can provide evidence for either "burst-phase" kinetics, which can reveal a slow product release step, or linear-phase kinetics, in which the chemical step can be rate-determining. With these assignments it becomes more straightforward to analyze the effects of mutations. We use examples from the hydroxysteroid dehydrogenases (AKR1Cs) and human steroid 5β-reductase (AKR1D1) to illustrate the utility of the approach, which are members of the aldo-keto reductase (AKR) superfamily.

  9. Explaining the atypical reaction profiles of heme enzymes with a novel mechanistic hypothesis and kinetic treatment.

    Directory of Open Access Journals (Sweden)

    Kelath Murali Manoj

    Full Text Available Many heme enzymes show remarkable versatility and atypical kinetics. The fungal extracellular enzyme chloroperoxidase (CPO characterizes a variety of one and two electron redox reactions in the presence of hydroperoxides. A structural counterpart, found in mammalian microsomal cytochrome P450 (CYP, uses molecular oxygen plus NADPH for the oxidative metabolism (predominantly hydroxylation of substrate in conjunction with a redox partner enzyme, cytochrome P450 reductase. In this study, we employ the two above-mentioned heme-thiolate proteins to probe the reaction kinetics and mechanism of heme enzymes. Hitherto, a substrate inhibition model based upon non-productive binding of substrate (two-site model was used to account for the inhibition of reaction at higher substrate concentrations for the CYP reaction systems. Herein, the observation of substrate inhibition is shown for both peroxide and final substrate in CPO catalyzed peroxidations. Further, analogy is drawn in the "steady state kinetics" of CPO and CYP reaction systems. New experimental observations and analyses indicate that a scheme of competing reactions (involving primary product with enzyme or other reaction components/intermediates is relevant in such complex reaction mixtures. The presence of non-selective reactive intermediate(s affords alternate reaction routes at various substrate/product concentrations, thereby leading to a lowered detectable concentration of "the product of interest" in the reaction milieu. Occam's razor favors the new hypothesis. With the new hypothesis as foundation, a new biphasic treatment to analyze the kinetics is put forth. We also introduce a key concept of "substrate concentration at maximum observed rate". The new treatment affords a more acceptable fit for observable experimental kinetic data of heme redox enzymes.

  10. Double perturbation series in the differential equations of enzyme kinetics

    Science.gov (United States)

    Fraser, Simon J.

    1998-07-01

    The connection between combined singular and ordinary perturbation methods and slow-manifold theory is discussed using the Michaelis-Menten model of enzyme catalysis as an example. This two-step mechanism is described by a planar system of ordinary differential equations (ODEs) with a fast transient and a slow "steady-state" decay mode. The systems of scaled nonlinear ODEs for this mechanism contain a singular (η) and an ordinary (ɛ) perturbation parameter: η multiplies the velocity component of the fast variable and dominates the fast-mode perturbation series; ɛ controls the decay toward equilibrium and dominates the slow-mode perturbation series. However, higher order terms in both series contain η and ɛ. Finite series expansions partially decouple the system of ODEs into fast-mode and slow-mode ODEs; infinite series expansions completely decouple these ODEs. Correspondingly, any slow-mode ODE approximately describes motion on M, the linelike slow manifold of the system, and in the infinite series limit this description is exact. Thus the perturbation treatment and the slow-manifold picture of the system are closely related. The functional equation for M is solved automatically with the manipulative language MAPLE. The formal η and ɛ single perturbation expansions for the slow mode yield the same double (η,ɛ) perturbation series expressions to given order. Generalizations of this procedure are discussed.

  11. Theory and simulation of diffusion-controlled Michaelis-Menten kinetics for a static enzyme in solution.

    Science.gov (United States)

    Park, Soohyung; Agmon, Noam

    2008-05-15

    We develop a uniform theory for the many-particle diffusion-control effects on the Michaelis-Menten scheme in solution, based on the Gopich-Szabo relaxation-time approximation (Gopich, I. V.; Szabo, A. J. Chem. Phys. 2002, 117, 507). We extend the many-particle simulation algorithm to the Michaelis-Menten case by utilizing the Green function previously derived for excited-state reversible geminate recombination with different lifetimes (Gopich, I. V.; Agmon, N. J. Chem. Phys. 2000, 110, 10433). Running the simulation for representative parameter sets in the time domain and under steady-state conditions, we find poor agreement with classical kinetics but excellent agreement with some of the modern theories for bimolecular diffusion-influenced reactions. Our simulation algorithm can be readily extended to the biologically interesting case of dense patches of membrane-bound enzymes.

  12. Testing Biochemistry Revisited : How In Vivo Metabolism Can Be Understood from In Vitro Enzyme Kinetics

    NARCIS (Netherlands)

    van Eunen, Karen; Kiewiet, Jose A. L.; Westerhoff, Hans V.; Bakker, Barbara M.

    2012-01-01

    A decade ago, a team of biochemists including two of us, modeled yeast glycolysis and showed that one of the most studied biochemical pathways could not be quite understood in terms of the kinetic properties of the constituent enzymes as measured in cell extract. Moreover, when the same model was la

  13. Appreciating Formal Similarities in the Kinetics of Homogeneous, Heterogeneous, and Enzyme Catalysis

    Science.gov (United States)

    Ashby, Michael T.

    2007-01-01

    Because interest in catalysts is widespread, the kinetics of catalytic reactions have been investigated by widely diverse groups of individuals, including chemists, engineers, and biologists. This has lead to redundancy in theories, particularly with regard to the topics of homogeneous, heterogeneous, and enzyme catalysis. From a pedagogical…

  14. Enzyme kinetics and identification of the rate-limiting step of enzymatic arabinoxylan degradation

    DEFF Research Database (Denmark)

    Rasmussen, Louise Enggaard; Xu, Cheng; Sørensen, Jens

    2012-01-01

    This study investigated the kinetics of multi-enzymatic degradation of soluble wheat arabinoxylan by monitoring the release of xylose and arabinose during designed treatments with mono-component enzymes at different substrate concentrations. The results of different combinations of α-l-arabinofur...

  15. Enzyme catalyzed oxidative gelation of sugar beet pectin: Kinetics and rheology

    DEFF Research Database (Denmark)

    Abang Zaidel, Dayang Norulfairuz; Chronakis, Ioannis S.; Meyer, Anne S.

    2012-01-01

    Sugar beet pectin (SBP) is a marginally utilized co-processing product from sugar production from sugar beets. In this study, the kinetics of oxidative gelation of SBP, taking place via enzyme catalyzed cross-linking of ferulic acid moieties (FA), was studied using small angle oscillatory measure...

  16. Estimation of the overall kinetic parameters of enzyme inactivation using an isoconversional method

    OpenAIRE

    2008-01-01

    Estimation of the overall kinetic parameters of enzyme inactivation using an isoconversional method correspondance: Corresponding author. Tel./fax: +40 213275116, +40 722624585 (Mobile). (Raducan, Adina) (Raducan, Adina) Department of Physical Chemistry--> , Faculty of Chemistry--> , University of Bucharest--> , Bd. Elisabeta 4-12--> , 030018--> , Bucharest--> - RO...

  17. Detailed enzyme kinetics in terms of biochemical species: study of citrate synthase.

    Directory of Open Access Journals (Sweden)

    Daniel A Beard

    Full Text Available The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant. The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data. This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

  18. Determining Enzyme Kinetics for Systems Biology with Nuclear Magnetic Resonance Spectroscopy

    Directory of Open Access Journals (Sweden)

    Johann J. Eicher

    2012-11-01

    Full Text Available Enzyme kinetics for systems biology should ideally yield information about the enzyme’s activity under in vivo conditions, including such reaction features as substrate cooperativity, reversibility and allostery, and be applicable to enzymatic reactions with multiple substrates. A large body of enzyme-kinetic data in the literature is based on the uni-substrate Michaelis–Menten equation, which makes unnatural assumptions about enzymatic reactions (e.g., irreversibility, and its application in systems biology models is therefore limited. To overcome this limitation, we have utilised NMR time-course data in a combined theoretical and experimental approach to parameterize the generic reversible Hill equation, which is capable of describing enzymatic reactions in terms of all the properties mentioned above and has fewer parameters than detailed mechanistic kinetic equations; these parameters are moreover defined operationally. Traditionally, enzyme kinetic data have been obtained from initial-rate studies, often using assays coupled to NAD(PH-producing or NAD(PH-consuming reactions. However, these assays are very labour-intensive, especially for detailed characterisation of multi-substrate reactions. We here present a cost-effective and relatively rapid method for obtaining enzyme-kinetic parameters from metabolite time-course data generated using NMR spectroscopy. The method requires fewer runs than traditional initial-rate studies and yields more information per experiment, as whole time-courses are analyzed and used for parameter fitting. Additionally, this approach allows real-time simultaneous quantification of all metabolites present in the assay system (including products and allosteric modifiers, which demonstrates the superiority of NMR over traditional spectrophotometric coupled enzyme assays. The methodology presented is applied to the elucidation of kinetic parameters for two coupled glycolytic enzymes from Escherichia coli

  19. Kinetics and spatial distribution of enzymes of carbon, nitrogen and phosphorus cycles in earthworm biopores

    Science.gov (United States)

    Hoang Thi Thu, Duyen; Razavi, Bahar S.

    2016-04-01

    Earthworms boost microbial activities and consequently form hotspots in soil. The distribution of enzyme activities inside the earthworm biopores is completely unknown. For the first time, we analyzed enzyme kinetics and visualized enzyme distribution inside and outside biopores by in situ soil zymography. Kinetic parameters (Vmax and Km) of 6 enzymes β-glucosidase (GLU), cellobiohydrolase (CBH), xylanase (XYL), chitinase (NAG), leucine aminopeptidase (LAP) and acid phosphatase (APT) were determined in biopores formed by Lumbricus terrestris L.. The spatial distributions of GLU, NAG and APT become visible via zymograms in comparison between earthworm-inhabited and earthworm-free soil. Zymography showed heterogeneous distribution of hotspots in the rhizosphere and biopores. The hotspot areas were 2.4 to 14 times larger in the biopores than in soil without earthworms. The significantly higher Vmax values for GLU, CBH, XYL, NAG and APT in biopores confirmed the stimulation of enzyme activities by earthworms. For CBH, XYL and NAG, the 2- to 3-fold higher Km values in biopores indicated different enzyme systems with lower substrate affinity compared to control soil. The positive effects of earthworms on Vmax were cancelled by the Km increase for CBH, XYL and NAG at a substrate concentration below 20 μmol g-1 soil. The change of enzyme systems reflected a shift in dominant microbial populations toward species with lower affinity to holo-celluloses and to N-acetylglucosamine, and with higher affinity to proteins as compared to the biopores-free soil. We conclude that earthworm biopores are microbial hotspots with much higher and dense distribution of enzyme activities compared to bulk soil. References Spohn M, Kuzyakov Y. (2014) Spatial and temporal dynamics of hotspots of enzyme activity in soil as affected by living and dead roots - a soil zymography analysis, Plant Soil 379: 67-77. Blagodatskaya, E., Kuzyakov, Y., 2013. Review paper: Active microorganisms in soil

  20. Kinetic expression for maltose production from soluble starch by simultaneous use of beta-amylase and debranching enzymes.

    Science.gov (United States)

    Shiraishi, F; Kawakami, K; Yuasa, A; Kojima, T; Kusunoki, K

    1987-08-20

    The kinetics of the hydrolysis of soluble starch by simultaneous use of beta-amylase and either isoamylase or pullulanse was studied experimentally for a wide range of subtrate and enzyme concentrations. A kinetic expression was constituted for maltose production by beta-armylase, which was stimulated by an increase in linear linkage portions due to the debranching enzyme on amylopectin molecules. As a result, calculations by the kinetic expression agreed with time course data under various conditions.

  1. Classical nucleation theory of homogeneous freezing of water: thermodynamic and kinetic parameters.

    Science.gov (United States)

    Ickes, Luisa; Welti, André; Hoose, Corinna; Lohmann, Ulrike

    2015-02-28

    The probability of homogeneous ice nucleation under a set of ambient conditions can be described by nucleation rates using the theoretical framework of Classical Nucleation Theory (CNT). This framework consists of kinetic and thermodynamic parameters, of which three are not well-defined (namely the interfacial tension between ice and water, the activation energy and the prefactor), so that any CNT-based parameterization of homogeneous ice formation is less well-constrained than desired for modeling applications. Different approaches to estimate the thermodynamic and kinetic parameters of CNT are reviewed in this paper and the sensitivity of the calculated nucleation rate to the choice of parameters is investigated. We show that nucleation rates are very sensitive to this choice. The sensitivity is governed by one parameter - the interfacial tension between ice and water, which determines the energetic barrier of the nucleation process. The calculated nucleation rate can differ by more than 25 orders of magnitude depending on the choice of parameterization for this parameter. The second most important parameter is the activation energy of the nucleation process. It can lead to a variation of 16 orders of magnitude. By estimating the nucleation rate from a collection of droplet freezing experiments from the literature, the dependence of these two parameters on temperature is narrowed down. It can be seen that the temperature behavior of these two parameters assumed in the literature does not match with the predicted nucleation rates from the fit in most cases. Moreover a comparison of all possible combinations of theoretical parameterizations of the dominant two free parameters shows that one combination fits the fitted nucleation rates best, which is a description of the interfacial tension coming from a molecular model [Reinhardt and Doye, J. Chem. Phys., 2013, 139, 096102] in combination with the activation energy derived from self-diffusion measurements [Zobrist

  2. A generalized cellular automata approach to modeling first order enzyme kinetics

    Indian Academy of Sciences (India)

    Abhishek Dutta; Saurajyoti Kar; Advait Apte; Ingmar Nopens; Denis Constales

    2015-04-01

    Biochemical processes occur through intermediate steps which are associated with the formation of reaction complexes. These enzyme-catalyzed biochemical reactions are inhibited in a number of ways such as inhibitors competing for the binding site directly, inhibitors deforming the allosteric site or inhibitors changing the structure of active substrate. Using an in silico approach, the concentration of various reaction agents can be monitored at every single time step, which are otherwise difficult to analyze experimentally. Cell-based models with discrete state variables, such as Cellular Automata (CA) provide an understanding of the organizational principles of interacting cellular systems to link the individual cell (microscopic) dynamics wit a particular collective (macroscopic) phenomenon. In this study, a CA model representing a first order enzyme kinetics with inhibitor activity is formulated. The framework of enzyme reaction rules described in this study is probabilistic. An extended von Neumann neighborhood with periodic boundary condition is implemented on a two-dimensional (2D) lattice framework. The effect of lattice-size variation is studied followed by a sensitivity analysis of the model output to the probabilistic parameters which represent various kinetic reaction constants in the enzyme kinetic model. This provides a deeper insight into the sensitivity of the CA model to these parameters. It is observed that cellular automata can capture the essential features of a discrete real system, consisting of space, time and state, structured with simple local rules without making complex implementations but resulting in complex but explainable patterns.

  3. Developing the enzyme-machine analogy: a non-mathematical approach to teaching Michaelis-Menten kinetics

    OpenAIRE

    Simon Brown

    2010-01-01

    The behavior of enzyme-catalyzed reactions is not made clear to many students by the standard mathematical description of enzyme kinetics. An enzyme-machine analogy is described that has made the details of the Michaelis-Menten mechanism and the associated kinetics more accessible with minimal use of mathematics. Students taught using the analogy appear to have fewer of the misconceptions than those taught using a more mathematical approach.

  4. Developing the enzyme-machine analogy: a non-mathematical approach to teaching Michaelis-Menten kinetics

    Directory of Open Access Journals (Sweden)

    Simon Brown

    2010-06-01

    Full Text Available The behavior of enzyme-catalyzed reactions is not made clear to many students by the standard mathematical description of enzyme kinetics. An enzyme-machine analogy is described that has made the details of the Michaelis-Menten mechanism and the associated kinetics more accessible with minimal use of mathematics. Students taught using the analogy appear to have fewer of the misconceptions than those taught using a more mathematical approach.

  5. Exact closed probability-free kinetic equation for system of classical particles with retarded interactions: towards microscopic foundation of kinetics

    CERN Document Server

    Zakharov, A Yu

    2016-01-01

    The exact closed equation of motion for microscopic distribution function of classical many-body system with account of interactions retardation between particles is derived. It is shown that interactions retardation leads to irreversible behaviour of many-body systems.

  6. Hepatic glucuronidation of resveratrol: interspecies comparison of enzyme kinetic profiles in human, mouse, rat, and dog.

    Science.gov (United States)

    Maier-Salamon, Alexandra; Böhmdorfer, Michaela; Thalhammer, Theresia; Szekeres, Thomas; Jaeger, Walter

    2011-01-01

    The enzyme kinetic profiles of the formation of resveratrol-3-O-glucuronide (R3G) and resveratrol-4'-O-glucuronide (R4'G) by liver microsomes from humans, dogs, and rodents were investigated. Glucuronidation by human and dog liver microsomes to R3G and R4'G occurred for about 65% of applied resveratrol, and was significantly reduced to 10% when substrate concentration was increased 10-fold. In contrast, rodent microsomes glucuronidated about 90% of applied resveratrol independently of substrate concentration. Furthermore, in mouse and rat liver microsomes, resveratrol was almost exclusively conjugated at position 3, whereas human and dog livers also glucuronidated resveratrol at position 4' (ratio R3G:R4'G = 5:1). Interspecies differences were also found when calculating the enzyme kinetic profiles of both conjugates. Formation of R4'G in human and dog microsomes followed Michaelis-Menten kinetics, while R3G showed substrate inhibition at higher resveratrol concentrations. In mouse and rat microsomes, however, both R3G and R4'G formation exhibited auto-activation kinetics. Formation of R3G and R4'G by recombinant UGT1A1 also showed substrate inhibition kinetics that led to decreased intrinsic clearance values, while UGT1A9-catalyzed glucuronidation demonstrated substrate inhibition kinetics at position 3 and Hill kinetics for the formation of R4'G. In conclusion, resveratrol glucuronidation exhibited species-dependent differences, with the dog as the animal model that most closely represents humans in terms of this process.

  7. Application of A Microfluidic Tool for the Determination of Enzyme Kinetics

    DEFF Research Database (Denmark)

    Ringborg, Rolf H.

    the limitations surround the reactor of a process,and with the performance of this being unknown, it is almost impossible to direct development. A focal point must therefore lie in the determination of kinetic models and howkinetic data can be obtained in a robust and generic way. Models for many enzymes...... in the collected retention time wavelength data. A major improvement over traditional techniques is the quantification of enzyme concentration and this makes it possible to use specific activities for model fitting. The setup takes advantage of microfluidic featuresand delivers semi‐automatic experimentation...

  8. A Numerical Procedure for Model Identifiability Analysis Applied to Enzyme Kinetics

    DEFF Research Database (Denmark)

    Daele, Timothy, Van; Van Hoey, Stijn; Gernaey, Krist;

    2015-01-01

    exercise, thereby bypassing the challenging task of model structure determination and identification. Parameter identification problems can thus lead to ill-calibrated models with low predictive power and large model uncertainty. Every calibration exercise should therefore be precededby a proper model...... and Pronzato (1997) and which can be easily set up for any type of model. In this paper the proposed approach is applied to the forward reaction rate of the enzyme kinetics proposed by Shin and Kim(1998). Structural identifiability analysis showed that no local structural model problems were occurring......The proper calibration of models describing enzyme kinetics can be quite challenging. In the literature, different procedures are available to calibrate these enzymatic models in an efficient way. However, in most cases the model structure is already decided on prior to the actual calibration...

  9. Kinetic and solvent deuterium isotope effects in the oxidation of putrescine catalysed by enzyme diamine oxidase.

    Science.gov (United States)

    Pałka, Katarzyna; Szymańska, Jolanta; Kańska, Marianna

    2013-01-01

    In this study, the kinetic isotope effects and solvent isotope effects in the reaction of the deamination of [(1R)-(2)H ] putrescine--catalysed by enzyme diamine oxidase (EC 1.4.3.6)--were determined using a non-competitive spectroscopic method. Putrescine, stereospecifically labelled with deuterium, was obtained by enzymatic decarboxylation of l-ornithine that was carried out in a fully deuteriated incubation medium.

  10. Measurement of enzyme kinetics and inhibitor constants using enthalpy arrays1

    OpenAIRE

    2009-01-01

    Enthalpy arrays enable label-free, solution-based calorimetric detection of molecular interactions in a 96-detector array format. Compared with conventional calorimetry, enthalpy arrays achieve a significant reduction of sample volume and measurement time through the combination of the small size of the detectors and ability to perform measurements in parallel. The current capabilities of the technology for studying enzyme-catalyzed reactions are demonstrated by determining the kinetic parame...

  11. Optimal design for goodness-of-fit of the Michaelis-Menten enzyme kinetic function

    OpenAIRE

    Wong, Weng Kee; Melas, Viatcheslav B.; Dette, Holger

    2004-01-01

    We construct efficient designs for the Michaelis-Menten enzyme kinetic model capable of checking model assumption. An extended model, called EMAX model is also considered for this purpose. This model is widely used in pharmacokinetics and reduces to the Michaelis- Menten model for a specific choice of the parameter setting. Our strategy is to find efficient designs for estimating the parameters in the EMAX model and at the same time test the validity of the Michaelis-Menten model against the ...

  12. Enzyme-catalyzed synthesis and kinetics of ultrasonic-assisted biodiesel production from waste tallow.

    Science.gov (United States)

    Adewale, Peter; Dumont, Marie-Josée; Ngadi, Michael

    2015-11-01

    The use of ultrasonic processing was evaluated for its ability to achieve adequate mixing while providing sufficient activation energy for the enzymatic transesterification of waste tallow. The effects of ultrasonic parameters (amplitude, cycle and pulse) and major reaction factors (molar ratio and enzyme concentration) on the reaction kinetics of biodiesel generation from waste tallow bio-catalyzed by immobilized lipase [Candida antarctica lipase B (CALB)] were investigated. Three sets of experiments namely A, B, and C were conducted. In experiment set A, two factors (ultrasonic amplitude and cycle) were investigated at three levels; in experiment set B, two factors (molar ratio and enzyme concentration) were examined at three levels; and in experiment set C, two factors (ultrasonic amplitude and reaction time) were investigated at five levels. A Ping Pong Bi Bi kinetic model approach was employed to study the effect of ultrasonic amplitude on the enzymatic transesterification. Kinetic constants of transesterification reaction were determined at different ultrasonic amplitudes (30%, 35%, 40%, 45%, and 50%) and enzyme concentrations (4, 6, and 8 wt.% of fat) at constant molar ratio (fat:methanol); 1:6, and ultrasonic cycle; 5 Hz. Optimal conditions for ultrasound-assisted biodiesel production from waste tallow were fat:methanol molar ratio, 1:4; catalyst level 6% (w/w of fat); reaction time, 20 min (30 times less than conventional batch processes); ultrasonic amplitude 40% at 5 Hz. The kinetic model results revealed interesting features of ultrasound assisted enzyme-catalyzed transesterification (as compared to conventional system): at ultrasonic amplitude 40%, the reaction activities within the system seemed to be steady after 20 min which means the reaction could proceed with or without ultrasonic mixing. Reversed phase high performance liquid chromatography indicated the biodiesel yield to be 85.6±0.08%.

  13. Mediated Electron Transfer at Redox Active Monolayers. Part 4: Kinetics of Redox Enzymes Coupled With Electron Mediators

    Directory of Open Access Journals (Sweden)

    Michael E.G. Lyons

    2003-01-01

    Full Text Available A detailed kinetic analysis of the pertinent physical processes underlying the operation of enzyme electrodes immobilized within alkane thiol self assembled monolayers is developed. These electrodes utilize a soluble mediator, which partitions into the monolayer, regenerates the active catalytic form of the enzyme and is re-oxidized at the underlying support electrode surface giving rise to a current which reflects kinetic events at the enzyme surface. Both the enzyme/substrate and enzyme mediator kinetics have been quantified fully in terms of a ping-pong mechanism for the former and Michaelis-Menten kinetics for the latter. The effect of substrate and mediator diffusion in solution have also been specifically considered and the latter processes have been shown to result in a complex expression for the reaction flux. Four limiting kinetic cases have been enumerated and simple expressions for the reaction flux in each of these rate limiting situations have been developed. Kinetic case diagrams have been presented as an aid to mechanistic diagnosis. The complicating effects of diffusive loss of reduced mediator from the enzyme layer have also been examined and the relation between the observed flux corresponding to reduced mediator oxidation at the support electrode and the substrate reaction flux in the enzyme layer have been quantified in terms of an efficiency factor. Results extracted from recently published practical realizations of immobilized monolayer enzyme systems have been discussed in the context of the proposed model analysis.

  14. KINETIC STUDY OF THE ENZYME CONVERSION OF STEAM EXPLODED PAULOWNIA TOMENTOSA TO GLUCOSE

    Directory of Open Access Journals (Sweden)

    Greta Radeva,

    2011-11-01

    Full Text Available Paulownia Tomentosa was pretreated by steam explosion. The cellulase complex NS 50013 and the β-glucosidase NS 50010 of Novozymes AS were used for the enzymatic conversion of cellulose to glucose. The kinetics of enzyme conversion was studied using the exponential kinetic equation valid for processes taking place at uniformly inhomogeneous surfaces. The kinetic coefficient of inhomogeneity accounts for the energy and entropy inhomogeneity of the system and depends on the temperature. It was established that both the activation energy and the pre-exponential factor increase simultaneously with increasing of conversion degree. A compensation effect between pre-exponential factor and activation energy was observed. The energetic hindrances established cannot be completely compensated by the positive effect of the pre-exponential factor increase. Hence, the activation energy has a determining influence on how quickly the rate of hydrolysis decreases.

  15. Production of lactobionic acid and sorbitol from lactose/fructose substrate using GFOR/GL enzymes from Zymomonas mobilis cells: a kinetic study.

    Science.gov (United States)

    Pedruzzi, Israel; da Silva, Eduardo A Borges; Rodrigues, Alírio E

    2011-07-10

    In this work, we have investigated the kinetics of the biotechnological production of lactobionic acid (LBA) and sorbitol by the catalytic action of glucose-fructose oxidoreductase (GFOR) and glucono-δ-lactonase (GL) enzymes. The cells of bacterium Zymomonas mobilis ATCC 29191 containing this enzymatic complex were submitted to permeabilization and reticulation procedures. The effect of the concentration of substrates on the rate of product formation using a mobilized cell system was investigated. The application of higher fructose concentration seems to not affect the initial rate of formation of the bionic acid. Under conditions of low initial concentration of lactose, the experimental kinetic data of the bi-substrate reaction were modelled by assuming a rate equation of the classical ping-pong mechanism. The found kinetic parameters displayed a low affinity of the GFOR enzyme for both substrates. The enzymatic system did not exhibit normal Michaelis-Menten kinetics in response to a change of concentration of lactose, when fructose was held constant, presenting a sigmoid relationship between initial velocity and substrate concentration. A rate equation based on Hill kinetics was used to describe the kinetic behaviour of this enzyme-substituted reaction at higher lactose concentrations. The results from batch experiments using immobilized cells within Ca-alginate beads revealed that there is no pronounced occurrence of mass transfer limitations on LBA production for beads with 1.2 mm in average diameter. This discussion aids for defining the best operating conditions to maximize the productivity for LBA and sorbitol in this bioconversion and also for reducing the complexity of downstream separation processes.

  16. Standardization of α-L-iduronidase enzyme assay with Michaelis-Menten kinetics.

    Science.gov (United States)

    Ou, Li; Herzog, Tyler L; Wilmot, Carrie M; Whitley, Chester B

    2014-02-01

    The lack of methodological uniformity in enzyme assays has been a long-standing difficulty, a problem for bench researchers, for the interpretation of clinical diagnostic tests, and an issue for investigational drug review. Illustrative of the problem, α-L-iduronidase enzyme catalytic activity is frequently measured with the substrate 4-methylumbelliferyl-α-L-iduronide (4MU-iduronide); however, final substrate concentrations used in different assays vary greatly, ranging from 25 μM to 1425 μM (Km ≈ 180 μM) making it difficult to compare results between laboratories. In this study, α-L-iduronidase was assayed with 15 different substrate concentrations. The resulting activity levels from the same specimens varied greatly with different substrate concentrations but, as a group, obeyed the expectations of Michaelis-Menten kinetics. Therefore, for the sake of improved comparability, it is proposed that α-L-iduronidase enzyme assays should be conducted either (1) under substrate saturating conditions; or (2) when concentrations are significantly below substrate saturation, with results standardized by arithmetic adjustment that considers Michaelis-Menten kinetics. The approach can be generalized to many other enzyme assays.

  17. About and beyond the Henri-Michaelis-Menten rate equation for single-substrate enzyme kinetics.

    Science.gov (United States)

    Bajzer, Zeljko; Strehler, Emanuel E

    2012-01-20

    For more than a century the simple single-substrate enzyme kinetics model and related Henri-Michaelis-Menten (HMM) rate equation have been thoroughly explored in various directions. In the present paper we are concerned with a possible generalization of this rate equation recently proposed by F. Kargi (BBRC 382 (2009) 157-159), which is assumed to be valid both in the case that the total substrate or enzyme is in excess and the quasi-steady-state is achieved. We demonstrate that this generalization is grossly inadequate and propose another generalization based on application of the quasi-steady-state condition and conservation equations for both enzyme and substrate. The standard HMM equation is derived by (a) assuming the quasi-steady-state condition, (b) applying the conservation equation only for the enzyme, and (c) assuming that the substrate concentration at quasi-steady-state can be approximated by the total substrate concentration [S](0). In our formula the rate is already expressed through [S](0), and we only assume that when quasi-steady-state is achieved the amount of product formed is negligible compared to [S](0). Numerical simulations show that our formula is generally more accurate than the HMM formula and also can provide a good approximation when the enzyme is in excess, which is not the case for the HMM formula. We show that the HMM formula can be derived from our expression by further assuming that the total enzyme concentration is negligible compared to [S](0).

  18. Renal histology before and after effective enzyme replacement therapy in a patient with classical Fabry's disease.

    Science.gov (United States)

    Hirashio, S; Taguchi, T; Naito, T; Maki, K; Ogata, S; Taniyama, K; Taniguchi, Y; Yorioka, N

    2009-05-01

    A 38-year-old man underwent renal biopsy because of proteinuria. It revealed swelling and vacuolation of glomerular epithelial cells, as well as myelin-like structures characteristic of Fabry's disease. Detection of decreased plasma activity of alpha-galactosidase A confirmed the diagnosis. Enzyme replacement therapy was provided with recombinant agalsidase-beta, resulting in improvement of his symptoms. When renal biopsy was repeated, specific staining for globotriaosylceramide showed that renal deposits were decreased by enzyme therapy.

  19. Evidence for communality in the primary determinants of CYP74 catalysis and of structural similarities between CYP74 and classical mammalian P450 enzymes.

    Science.gov (United States)

    Hughes, Richard K; Yousafzai, Faridoon K; Ashton, Ruth; Chechetkin, Ivan R; Fairhurst, Shirley A; Hamberg, Mats; Casey, Rod

    2008-09-01

    In silico structural analysis of CYP74C3, a membrane-associated P450 enzyme from the plant Medicago truncatula (barrel medic) with hydroperoxide lyase (HPL) specificity, showed that it had strong similarities to the structural folds of the classical microsomal P450 enzyme from rabbits (CYP2C5). It was not only the secondary structure predictions that supported the analysis but site directed mutagenesis of the substrate interacting residues was also consistent with it. This led us to develop a substrate-binding model of CYP74C3 which predicted three amino acid residues, N285, F287, and G288 located in the putative I-helix and distal haem pocket of CYP74C3 to be in close proximity to the preferred substrate 13-HPOTE. These residues were judged to be in equivalent positions to those identified in SRS-4 of CYP2C5. Significance of the residues and their relevance to the model were further assessed by site directed mutagenesis of the three residues followed by EPR spectroscopic and detailed kinetic investigations of the mutated proteins in the presence and absence of detergent. Although point mutation of the residues had no effect on the haem content of the mutated proteins, significant effects on the spin state equilibrium of the haem iron were noted. Kinetic effects of the mutations, which were investigated using three different substrates, were dramatic in nature. In the presence of detergent with the preferred substrate (13-HPOTE), the catalytic center activities and substrate binding affinities of the mutant proteins were reduced by a factor of 8-32 and 4-12, respectively, compared with wild-type--a two orders of magnitude reduction in catalytic efficiencies. We believe this is the first report where primary determinants of catalysis for any CYP74 enzyme, which are fully consistent with our model, have been identified. Our working model predicts that N285 is close enough to suggest that a hydrogen bond with the peroxy group of the enzyme substrate 13-HPOTE is

  20. An Integrated Circuit for Chip-Based Analysis of Enzyme Kinetics and Metabolite Quantification.

    Science.gov (United States)

    Cheah, Boon Chong; Macdonald, Alasdair Iain; Martin, Christopher; Streklas, Angelos J; Campbell, Gordon; Al-Rawhani, Mohammed A; Nemeth, Balazs; Grant, James P; Barrett, Michael P; Cumming, David R S

    2016-06-01

    We have created a novel chip-based diagnostic tools based upon quantification of metabolites using enzymes specific for their chemical conversion. Using this device we show for the first time that a solid-state circuit can be used to measure enzyme kinetics and calculate the Michaelis-Menten constant. Substrate concentration dependency of enzyme reaction rates is central to this aim. Ion-sensitive field effect transistors (ISFET) are excellent transducers for biosensing applications that are reliant upon enzyme assays, especially since they can be fabricated using mainstream microelectronics technology to ensure low unit cost, mass-manufacture, scaling to make many sensors and straightforward miniaturisation for use in point-of-care devices. Here, we describe an integrated ISFET array comprising 2(16) sensors. The device was fabricated with a complementary metal oxide semiconductor (CMOS) process. Unlike traditional CMOS ISFET sensors that use the Si3N4 passivation of the foundry for ion detection, the device reported here was processed with a layer of Ta2O5 that increased the detection sensitivity to 45 mV/pH unit at the sensor readout. The drift was reduced to 0.8 mV/hour with a linear pH response between pH 2-12. A high-speed instrumentation system capable of acquiring nearly 500 fps was developed to stream out the data. The device was then used to measure glucose concentration through the activity of hexokinase in the range of 0.05 mM-231 mM, encompassing glucose's physiological range in blood. Localised and temporal enzyme kinetics of hexokinase was studied in detail. These results present a roadmap towards a viable personal metabolome machine.

  1. Nonlinear temperature sensitivity of enzyme kinetics explains canceling effect - a case study on loamy haplic Luvisol

    Directory of Open Access Journals (Sweden)

    Bahar S. eRazavi

    2015-10-01

    Full Text Available The temperature sensitivity of enzymes responsible for organic matter decomposition in soil is crucial for predicting the effects of global warming on the carbon cycle and sequestration. We tested the hypothesis that differences in temperature sensitivity of enzyme kinetic parameters Vmax and Km will lead to a canceling effect: strong reduction of temperature response of catalytic reactions. Short-term temperature response of Vmax and Km of three hydrolytic enzymes responsible for decomposition of cellulose (β-glucosidase, cellobiohydrolase and hemicelluloses (xylanase were analyzed in situ from 0 to 40 °C. The apparent activation energy varied between enzymes from 20.7 to 35.2 kJ mol-1 corresponding to the Q10 values of the enzyme activities of 1.4–1.9 (with Vmax-Q10 1.0–2.5 and Km-Q10 0.94–2.3. Temperature response of all tested enzymes fitted well to the Arrhenius equation. Despite that, the fitting of Arrhenius model revealed the non-linear increase of two cellulolytic enzymes activities with two distinct thresholds at 10–15 °C and 25–30 °C, which were less pronounced for xylanase. The nonlinearity between 10 and 15 °C was explained by 30–80% increase in Vmax. At 25–30 °C, however, the abrupt decrease of enzyme-substrate affinity was responsible for non-linear increase of enzyme activities. Our study is the first demonstrating nonlinear response of Vmax and Km to temperature causing canceling effect, which was most strongly pronounced at low substrate concentrations and at temperatures above 15 °C. Under cold climate, however, the regulation of hydrolytic activity by canceling in response to warming is negligible because canceling was never observed below 10 °C. The canceling, therefore, can be considered as natural mechanism reducing the effects of global warming on decomposition of soil organics at moderate temperatures. The non-linearity of enzyme responses to warming and the respective thresholds should therefore be

  2. A new multi-wavelength model-based method for determination of enzyme kinetic parameters.

    Science.gov (United States)

    Sorouraddin, Mohammad-Hossein; Amini, Kaveh; Naseri, Abdolhossein; Vallipour, Javad; Hanaee, Jalal; Rashidi, Mohammad-Reza

    2010-09-01

    Lineweaver-Burk plot analysis is the most widely used method to determine enzyme kinetic parameters. In the spectrophotometric determination of enzyme activity using the Lineweaver-Burk plot, it is necessary to find a wavelength at which only the substrate or the product has absorbance without any spectroscopic interference of the other reaction components. Moreover, in this method, different initial concentrations of the substrate should be used to obtain the initial velocities required for Lineweaver-Burk plot analysis. In the present work, a multi-wavelength model-based method has been developed and validated to determine Michaelis-Menten constants for some enzyme reactions. In this method, a selective wavelength region and several experiments with different initial concentrations of the substrate are not required. The absorbance data of the kinetic assays are fitted by non-linear regression coupled to the numeric integration of the related differential equation. To indicate the applicability of the proposed method, the Michaelis-Menten constants for the oxidation of phenanthridine, 6-deoxypenciclovir and xanthine by molybdenum hydroxylases were determined using only a single initial concentration of the substrate, regardless of any spectral overlap.

  3. Purification, kinetic behavior, and regulation of NAD(P)+ malic enzyme of tumor mitochondria.

    Science.gov (United States)

    Moreadith, R W; Lehninger, A L

    1984-05-25

    The purification and kinetic characterization of an NAD(P)+-malic enzyme from 22aH mouse hepatoma mitochondria are described. The enzyme was purified 328-fold with a final yield of 51% and specific activity of 38.1 units/mg of protein by employing DEAE-cellulose chromatography and an ATP affinity column. Sephadex G-200 chromatography yielded a native Mr = 240,000. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a major subunit with Mr = 61,000, suggesting a tetrameric structure, and also showed that the preparation contained less than 10% polypeptide impurities. Use of the ATP affinity column required the presence of MnCl2 and fumarate (an allosteric activator) in the elution buffers. In the absence of fumarate, the Michaelis constants for malate, NAD+, and NADP+ were 3.6 mM, 55 microM, and 72 microM, respectively; in the presence of fumarate (2 mM), the constants were 0.34 mM, 9 microM, and 13 microM, respectively. ATP was shown to be an allosteric inhibitor, competitive with malate. However, the inhibition by ATP displayed hyperbolic competitive kinetics with a KI (ATP) of 80 microM (minus fumarate) and 0.5 mM (plus 2 mM fumarate). The allosteric properties of the enzyme are integrated into a rationale for its specific role in the pathways of malate and glutamate oxidation in tumor mitochondria.

  4. A new multi-wavelength model-based method for determination of enzyme kinetic parameters

    Indian Academy of Sciences (India)

    Mohammad-Hossein Sorouraddin; Kaveh Amini; Abdolhossein Naseri; Javad Vallipour; Jalal Hanaee; Mohammad-Reza Rashidi

    2010-09-01

    Lineweaver–Burk plot analysis is the most widely used method to determine enzyme kinetic parameters. In the spectrophotometric determination of enzyme activity using the Lineweaver–Burk plot, it is necessary to find a wavelength at which only the substrate or the product has absorbance without any spectroscopic interference of the other reaction components. Moreover, in this method, different initial concentrations of the substrate should be used to obtain the initial velocities required for Lineweaver–Burk plot analysis. In the present work, a multi-wavelength model-based method has been developed and validated to determine Michaelis–Menten constants for some enzyme reactions. In this method, a selective wavelength region and several experiments with different initial concentrations of the substrate are not required. The absorbance data of the kinetic assays are fitted by non-linear regression coupled to the numeric integration of the related differential equation. To indicate the applicability of the proposed method, the Michaelis–Menten constants for the oxidation of phenanthridine, 6-deoxypenciclovir and xanthine by molybdenum hydroxylases were determined using only a single initial concentration of the substrate, regardless of any spectral overlap.

  5. Epoxide hydrolase-catalyzed enantioselective conversion of trans-stilbene oxide: Insights into the reaction mechanism from steady-state and pre-steady-state enzyme kinetics.

    Science.gov (United States)

    Archelas, Alain; Zhao, Wei; Faure, Bruno; Iacazio, Gilles; Kotik, Michael

    2016-02-01

    A detailed kinetic study based on steady-state and pre-steady-state measurements is described for the highly enantioselective epoxide hydrolase Kau2. The enzyme, which is a member of the α/β-hydrolase fold family, preferentially reacts with the (S,S)-enantiomer of trans-stilbene oxide (TSO) with an E value of ∼200. The enzyme follows a classical two-step catalytic mechanism with formation of an alkyl-enzyme intermediate in the first step and hydrolysis of this intermediate in a rate-limiting second step. Tryptophan fluorescence quenching during TSO conversion appears to correlate with alkylation of the enzyme. The steady-state data are consistent with (S,S) and (R,R)-TSO being two competing substrates with marked differences in k(cat) and K(M) values. The high enantiopreference of the epoxide hydrolase is best explained by pronounced differences in the second-order alkylation rate constant (k2/K(S)) and the alkyl-enzyme hydrolysis rate k3 between the (S,S) and (R,R)-enantiomers of TSO. Our data suggest that during conversion of (S,S)-TSO the two active site tyrosines, Tyr(157) and Tyr(259), serve mainly as electrophilic catalysts in the alkylation half-reaction, polarizing the oxirane oxygen of the bound epoxide through hydrogen bond formation, however, without fully donating their hydrogens to the forming alkyl-enzyme intermediate.

  6. Enzyme Kinetics by Directly Imaging a Porous Silicon Microfluidic Reactor Using Desorption/Ionization on Silicon Mass Spectrometry

    NARCIS (Netherlands)

    Nichols, Kevin P.; Azoz, Seyla; Gardeniers, Han J.G.E.

    2008-01-01

    Enzyme kinetics were obtained in a porous silicon microfluidic channel by combining an enzyme and substrate droplet, allowing them to react and deposit a small amount of residue on the channel walls, and then analyzing this residue by directly ionizing the channel walls using a matrix assisted laser

  7. Large-scale ruthenium- and enzyme-catalyzed dynamic kinetic resolution of (rac-1-phenylethanol

    Directory of Open Access Journals (Sweden)

    Bäckvall Jan-E

    2007-12-01

    Full Text Available Abstract The scale-up of the ruthenium- and enzyme-catalyzed dynamic kinetic resolution (DKR of (rac-1-phenylethanol (2 is addressed. The immobilized lipase Candida antarctica lipase B (CALB was employed for the resolution, which shows high enantioselectivity in the transesterification. The ruthenium catalyst used, (η 5-C5Ph5RuCl(CO2 1, was shown to possess very high reactivity in the "in situ" redox racemization of 1-phenylethanol (2 in the presence of the immobilized enzyme, and could be used in 0.05 mol% with high efficiency. Commercially available isopropenyl acetate was employed as acylating agent in the lipase-catalyzed transesterifications, which makes the purification of the product very easy. In a successful large-scale DKR of 2, with 0.05 mol% of 1, (R-1-phenylethanol acetate (3 was obtained in 159 g (97% yield in excellent enantiomeric excess (99.8% ee.

  8. Enzyme kinetics and transport in a system crowded by mobile macromolecules.

    Science.gov (United States)

    Echeverria, Carlos; Kapral, Raymond

    2015-11-21

    The dynamics of an elastic network model for the enzyme 4-oxalocrotonate tautomerase is studied in a system crowded by mobile macromolecules, also modeled by elastic networks. The system includes a large number of solvent molecules, as well as substrate and product molecules which undergo catalytic reactions with this hexameric protein. The time evolution of the entire system takes place through a hybrid dynamics that combines molecular dynamics for solute species and multiparticle collision dynamics for the solvent. It is shown that crowding leads to subdiffusive dynamics for the protein, in accord with many studies of diffusion in crowded environments, and increases orientational relaxation times. The enzyme reaction kinetics is also modified by crowding. The effective Michaelis constant decreases with crowding volume fraction, and this decrease is attributed to excluded volume effects, which dominate over effects due to reduced substrate diffusion that would cause the Michaelis constant to increase.

  9. Refolding and enzyme kinetic studies on the ferrochelatase of the cyanobacterium Synechocystis sp. PCC 6803.

    Directory of Open Access Journals (Sweden)

    Patrik Storm

    Full Text Available Heme is a cofactor for proteins participating in many important cellular processes, including respiration, oxygen metabolism and oxygen binding. The key enzyme in the heme biosynthesis pathway is ferrochelatase (protohaem ferrolyase, EC 4.99.1.1, which catalyzes the insertion of ferrous iron into protoporphyrin IX. In higher plants, the ferrochelatase enzyme is localized not only in mitochondria, but also in chloroplasts. The plastidic type II ferrochelatase contains a C-terminal chlorophyll a/b (CAB motif, a conserved hydrophobic stretch homologous to the CAB domain of plant light harvesting proteins and light-harvesting like proteins. This type II ferrochelatase, found in all photosynthetic organisms, is presumed to have evolved from the cyanobacterial ferrochelatase. Here we describe a detailed enzymological study on recombinant, refolded and functionally active type II ferrochelatase (FeCh from the cyanobacterium Synechocystis sp. PCC 6803. A protocol was developed for the functional refolding and purification of the recombinant enzyme from inclusion bodies, without truncation products or soluble aggregates. The refolded FeCh is active in its monomeric form, however, addition of an N-terminal His(6-tag has significant effects on its enzyme kinetics. Strikingly, removal of the C-terminal CAB-domain led to a greatly increased turnover number, k(cat, compared to the full length protein. While pigments isolated from photosynthetic membranes decrease the activity of FeCh, direct pigment binding to the CAB domain of FeCh was not evident.

  10. Refolding and enzyme kinetic studies on the ferrochelatase of the cyanobacterium Synechocystis sp. PCC 6803.

    Science.gov (United States)

    Storm, Patrik; Tibiletti, Tania; Hall, Michael; Funk, Christiane

    2013-01-01

    Heme is a cofactor for proteins participating in many important cellular processes, including respiration, oxygen metabolism and oxygen binding. The key enzyme in the heme biosynthesis pathway is ferrochelatase (protohaem ferrolyase, EC 4.99.1.1), which catalyzes the insertion of ferrous iron into protoporphyrin IX. In higher plants, the ferrochelatase enzyme is localized not only in mitochondria, but also in chloroplasts. The plastidic type II ferrochelatase contains a C-terminal chlorophyll a/b (CAB) motif, a conserved hydrophobic stretch homologous to the CAB domain of plant light harvesting proteins and light-harvesting like proteins. This type II ferrochelatase, found in all photosynthetic organisms, is presumed to have evolved from the cyanobacterial ferrochelatase. Here we describe a detailed enzymological study on recombinant, refolded and functionally active type II ferrochelatase (FeCh) from the cyanobacterium Synechocystis sp. PCC 6803. A protocol was developed for the functional refolding and purification of the recombinant enzyme from inclusion bodies, without truncation products or soluble aggregates. The refolded FeCh is active in its monomeric form, however, addition of an N-terminal His(6)-tag has significant effects on its enzyme kinetics. Strikingly, removal of the C-terminal CAB-domain led to a greatly increased turnover number, k(cat), compared to the full length protein. While pigments isolated from photosynthetic membranes decrease the activity of FeCh, direct pigment binding to the CAB domain of FeCh was not evident.

  11. Kinetic model for the co-action of beta-amylase and debranching enzymes in the production of maltose.

    Science.gov (United States)

    Jiahua, Z

    1999-03-05

    The kinetics of the hydrolysis of starch with beta-amylase and debranching enzymes was studied. The hydrolysis of the alpha-1, 6-glycoside bonds of the substrate by debranching enzymes does not create any new nonreducing ends, so debranching enzyme promotes the action of beta-amylase not by increasing the concentration of the substrate of beta-amylase but by increasing the linear linkage portion of the substrate. The introduction of an effective chain length function was used to formulate a kinetic model.

  12. Another Look at the Mechanisms of Hydride Transfer Enzymes from Quantum and Classical Transition Path Sampling

    Science.gov (United States)

    Dzierlenga, Michael; Antoniou, Dimitri; Schwartz, Steven

    2015-03-01

    The mechanisms involved in enzymatic hydride transfer have been studies for years but questions remain, due to the difficulty in determining the participation of protein dynamics and quantum effects, especially hydrogen tunneling. In this study, we use transition path sampling (TPS) with normal mode centroid molecular dynamics (CMD) to calculate the barrier to hydride transfer in yeast alcohol dehydrogenase (YADH) and lactate dehydrogenase (LDH). Calculation of the work applied to the hydride during the reaction allows for observation of the change in barrier height due to inclusion of quantum effects. Additionally, the same calculations were performed using deuterium as the transferring particle to validate our methods with experimentally measured kinetic isotope effects. The change in barrier height in YADH upon inclusion of quantum effects is indicative of a zero-point energy contribution, and is evidence that the protein mediates a near-barrierless transfer of the rate-limiting hydride. Calculation of kinetic isotope effects using the average difference in barrier between hydride and deuteride agreed well with experimental results. The authors acknowledge the support of the National Institutes of Health Grants GM068036 and GM102226.

  13. Enzyme kinetic measurements using a droplet-based microfluidic system with a concentration gradient.

    Science.gov (United States)

    Bui, Minh-Phuong Ngoc; Li, Cheng Ai; Han, Kwi Nam; Choo, Jaebum; Lee, Eun Kyu; Seong, Gi Hun

    2011-03-01

    In this paper, we propose a microfluidic device that is capable of generating a concentration gradient followed by parallel droplet formation within channels with a simple T-junction geometry. Linear concentration gradient profiles can be obtained based on fluid diffusion under laminar flow. Optimized conditions for generating a linear concentration gradient and parallel droplet formation were investigated using fluorescent dye. The concentration gradient profile under diffusive mixing was dominated by the flow rate at sample inlets, while parallel droplet formation was affected by the channel geometry at both the inlet and outlet. The microfluidic device was experimentally characterized using optimal layout and operating conditions selected through a design process. Furthermore, in situ enzyme kinetic measurements of the β-galactosidase-catalyzed hydrolysis of resorufin-β-d-galactopyranoside were performed to demonstrate the application potential of our simple, time-effective, and low sample volume microfluidic device. We expect that, in addition to enzyme kinetics, drug screening and clinical diagnostic tests can be rapidly and accurately performed using this droplet-based microfluidic system.

  14. Microbial respiration and kinetics of extracellular enzymes activities through rhizosphere and detritusphere at agricultural site

    Science.gov (United States)

    Löppmann, Sebastian; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2014-05-01

    Rhizosphere and detritusphere are soil microsites with very high resource availability for microorganisms affecting their biomass, composition and functions. In the rhizosphere low molecular compounds occur with root exudates and low available polymeric compounds, as belowground plant senescence. In detritusphere the substrate for decomposition is mainly a polymeric material of low availability. We hypothesized that microorganisms adapted to contrasting quality and availability of substrates in the rhizosphere and detritusphere are strongly different in affinity of hydrolytic enzymes responsible for decomposition of organic compounds. According to common ecological principles easily available substrates are quickly consumed by microorganisms with enzymes of low substrate affinity (i.e. r-strategists). The slow-growing K-strategists with enzymes of high substrate affinity are better adapted for growth on substrates of low availability. Estimation of affinity of enzyme systems to the substrate is based on Michaelis-Menten kinetics, reflecting the dependency of decomposition rates on substrate amount. As enzymes-mediated reactions are substrate-dependent, we further hypothesized that the largest differences in hydrolytic activity between the rhizosphere and detritusphere occur at substrate saturation and that these differences are smoothed with increasing limitation of substrate. Affected by substrate limitation, microbial species follow a certain adaptation strategy. To achieve different depth gradients of substrate availability 12 plots on an agricultural field were established in the north-west of Göttingen, Germany: 1) 4 plots planted with maize, reflecting lower substrate availability with depth; 2) 4 unplanted plots with maize litter input (0.8 kg m-2 dry maize residues), corresponding to detritusphere; 3) 4 bare fallow plots as control. Maize litter was grubbed homogenously into the soil at the first 5 cm to ensure comparable conditions for the herbivore and

  15. Another Look at the Mechanisms of Hydride Transfer Enzymes with Quantum and Classical Transition Path Sampling.

    Science.gov (United States)

    Dzierlenga, Michael W; Antoniou, Dimitri; Schwartz, Steven D

    2015-04-02

    The mechanisms involved in enzymatic hydride transfer have been studied for years, but questions remain due, in part, to the difficulty of probing the effects of protein motion and hydrogen tunneling. In this study, we use transition path sampling (TPS) with normal mode centroid molecular dynamics (CMD) to calculate the barrier to hydride transfer in yeast alcohol dehydrogenase (YADH) and human heart lactate dehydrogenase (LDH). Calculation of the work applied to the hydride allowed for observation of the change in barrier height upon inclusion of quantum dynamics. Similar calculations were performed using deuterium as the transferring particle in order to approximate kinetic isotope effects (KIEs). The change in barrier height in YADH is indicative of a zero-point energy (ZPE) contribution and is evidence that catalysis occurs via a protein compression that mediates a near-barrierless hydride transfer. Calculation of the KIE using the difference in barrier height between the hydride and deuteride agreed well with experimental results.

  16. Kinetically influenced terms for solute transport affected by heterogeneous and homogeneous classical reactions

    Science.gov (United States)

    Bahr, J.M.

    1990-01-01

    This paper extends a four-step derivation procedure, previously presented for cases of transport affected by surface reactions, to transport problems involving homogeneous reactions. Derivations for these classes of reactions are used to illustrate the manner in which mathematical differences between reaction classes are reflected in the mathematical derivation procedures required to identify kinetically influenced terms. Simulation results for a case of transport affected by a single solution phase complexation reaction and for a case of transport affected by a precipitation-dissolution reaction are used to demonstrate the nature of departures from equilibrium-controlled transport as well as the use of kinetically influenced terms in determining criteria for the applicability of the local equilibrium assumption. A final derivation for a multireaction problem demonstrates the application of the generalized procedure to a case of transport affected by reactions of several classes. -from Author

  17. Sequence diversity of NanA manifests in distinct enzyme kinetics and inhibitor susceptibility

    Science.gov (United States)

    Xu, Zhongli; von Grafenstein, Susanne; Walther, Elisabeth; Fuchs, Julian E.; Liedl, Klaus R.; Sauerbrei, Andreas; Schmidtke, Michaela

    2016-04-01

    Streptococcus pneumoniae is the leading pathogen causing bacterial pneumonia and meningitis. Its surface-associated virulence factor neuraminidase A (NanA) promotes the bacterial colonization by removing the terminal sialyl residues from glycoconjugates on eukaryotic cell surface. The predominant role of NanA in the pathogenesis of pneumococci renders it an attractive target for therapeutic intervention. Despite the highly conserved activity of NanA, our alignment of the 11 NanAs revealed the evolutionary diversity of this enzyme. The amino acid substitutions we identified, particularly those in the lectin domain and in the insertion domain next to the catalytic centre triggered our special interest. We synthesised the representative NanAs and the mutagenized derivatives from E. coli for enzyme kinetics study and neuraminidase inhibitor susceptibility test. Via molecular docking we got a deeper insight into the differences between the two major variants of NanA and their influence on the ligand-target interactions. In addition, our molecular dynamics simulations revealed a prominent intrinsic flexibility of the linker between the active site and the insertion domain, which influences the inhibitor binding. Our findings for the first time associated the primary sequence diversity of NanA with the biochemical properties of the enzyme and with the inhibitory efficiency of neuraminidase inhibitors.

  18. Estimation of Kinetic Parameters for Enzyme Catalysed Batch Bioreactor for the Production of Ethanol from Corn

    Directory of Open Access Journals (Sweden)

    Z. R. Yelebe

    2014-03-01

    Full Text Available This paper addresses the challenge of estimating various kinetic parameters for the design of an optimized enzyme catalysed batch bioreactor of high efficiency and yield. Mathematical models were developed to describe the batch reaction time in relation to the substrate, enzyme and product concentration. The results obtained from the plots generated were: 35.50gmol/l.hr for the velocity of reaction of the enzymes (Vmax, 0.10049hr-1 for the maximum specific growth rate (µmax 826.45gmol/l for the Michaelis-Menten constant (Km, 0.005402577 for maintenance coefficient (Ms, 10.104kgCx/kgCs for yield of cell weight per unit weight of substrate (Ycx/CS, 0.05436kgCp/kgCs for yield of product weight per unit weight of substrate utilized (Ycp/CS and 0.01416 for endogenous decay coefficient (Kd for the design of the batch biochemical reactor. Hence, they are useful parameters for predicting the most appropriate batch reaction conditions and the efficiency of the bioreactor. The mathematical model predictions showed that it can be considered as a good complimentary tool to real system since the simulation results of the mathematical model agrees with experimental data reported in literature.

  19. Some Investigations on Protease Enzyme Production Kinetics Using Bacillus licheniformis BBRC 100053 and Effects of Inhibitors on Protease Activity

    Directory of Open Access Journals (Sweden)

    Zahra Ghobadi Nejad

    2014-01-01

    Full Text Available Due to great commercial application of protease, it is necessary to study kinetic characterization of this enzyme in order to improve design of enzymatic reactors. In this study, mathematical modeling of protease enzyme production kinetics which is derived from Bacillus licheniformis BBRC 100053 was studied (at 37°C, pH 10 after 73 h in stationary phase, and 150 rpm. The aim of the present paper was to determine the best kinetic model and kinetic parameters for production of protease and calculating Ki (inhibition constant of different inhibitors to find the most effective one. The kinetic parameters Km (Michaelis-Menten constant and Vm (maximum rate were calculated 0.626 mM and 0.0523 mM/min. According to the experimental results, using DFP (diisopropyl fluorophosphate and PMSF (phenylmethanesulfonyl fluoride as inhibitors almost 50% of the enzyme activity could be inhibited when their concentrations were 0.525 and 0.541 mM, respectively. Ki for DFP and PMSF were 0.46 and 0.56 mM, respectively. Kinetic analysis showed that the Lineweaver-Burk model was the best fitting model for protease production kinetics DFP was more effective than PMSF and both of them should be covered in the group of noncompetitive inhibitors.

  20. Kinematic and kinetic analysis of the fouetté turn in classical ballet.

    Science.gov (United States)

    Imura, Akiko; Iino, Yoichi; Kojima, Takeji

    2010-11-01

    The fouetté turn in classical ballet dancing is a continuous turn with the whipping of the gesture leg and the arms and the bending and stretching of the supporting leg. The knowledge of the movement intensities of both legs for the turn would be favorable for the conditioning of the dancer's body. The purpose of this study was to estimate the intensities. The hypothesis of this study was that the intensities were higher in the supporting leg than in the gesture leg. The joint torques of both legs were determined in the turns performed by seven experienced female classical ballet dancers with inverse dynamics using three high-speed cine cameras and a force platform. The hip abductor torque, knee extensor and plantar flexor torques of the supporting leg were estimated to be exerted up to their maximum levels and the peaks of the torques were larger than the peaks of their matching torques of the gesture leg. Thus, the hypothesis was partly supported. Training of the supporting leg rather than the gesture leg would help ballet dancers perform many revolutions of the fouetté turn continuously.

  1. Exact and approximate solutions for a century-old problem: A general treatment of Henri-Michaelis-Menten enzyme kinetics

    OpenAIRE

    2009-01-01

    A different view of Henri-Michaelis-Menten (HMM) enzyme kinetics is presented. In the first part of the paper, a simplified but useful description that stresses the cyclic nature of the catalytic process is introduced. The time-dependence of the substrate concentration after the initial transient phase is derived in a simple way that dispenses the mathematical technique known as quasi-steady-state approximation. In the second part of the paper an exact one-dimensional formulation of HMM kinet...

  2. Compartmental and enzyme kinetic modeling to elucidate the biotransformation pathway of a centrally acting antitrypanosomal prodrug.

    Science.gov (United States)

    Generaux, Claudia N; Ainslie, Garrett R; Bridges, Arlene S; Ismail, Mohamed A; Boykin, David W; Tidwell, Richard R; Thakker, Dhiren R; Paine, Mary F

    2013-02-01

    DB868 [2,5-bis [5-(N-methoxyamidino)-2-pyridyl] furan], a prodrug of the diamidine DB829 [2,5-bis(5-amidino-2-pyridyl) furan], has demonstrated efficacy in murine models of human African trypanosomiasis. A cross-species evaluation of prodrug bioconversion to the active drug is required to predict the disposition of prodrug, metabolites, and active drug in humans. The phase I biotransformation of DB868 was elucidated using liver microsomes and sandwich-cultured hepatocytes from humans and rats. All systems produced four NADPH-dependent metabolites via O-demethylation (M1, M2) and N-dehydroxylation (M3, M4). Compartmental kinetic modeling of the DB868 metabolic pathway suggested an unusual N-demethoxylation reaction that was supported experimentally. A unienzyme Michaelis-Menten model described the kinetics of M1 formation by human liver microsomes (HLMs) (K(m), 11 μM; V(max), 340 pmol/min/mg), whereas a two-enzyme model described the kinetics of M1 formation by rat liver microsomes (RLMs) (K(m1), 0.5 μM; V(max1), 12 pmol/min/mg; K(m2), 27 μM; V(max2), 70 pmol/min/mg). Human recombinant CYP1A2, CYP3A4, and CYP4F2, rat recombinant Cyp1a2 and Cyp2d2, and rat purified Cyp4f1 catalyzed M1 formation. M2 formation by HLMs exhibited allosteric kinetics (S(50), 18 μM; V(max), 180 pmol/mg), whereas M2 formation by RLMs was negligible. Recombinant CYP1A2/Cyp1a2 catalyzed M2 formation. DB829 was detected in trace amounts in HLMs at the end of the 180-min incubation and was detected readily in sandwich-cultured hepatocytes from both species throughout the 24-h incubation. These studies demonstrated that DB868 biotransformation to DB829 is conserved between humans and rats. An improved understanding of species differences in the kinetics of DB829 formation would facilitate preclinical development of a promising antitrypanosomal prodrug.

  3. Kinetic substrate quantification by fitting the enzyme reaction curve to the integrated Michaelis-Menten equation.

    Science.gov (United States)

    Liao, Fei; Tian, Kao-Cong; Yang, Xiao; Zhou, Qi-Xin; Zeng, Zhao-Chun; Zuo, Yu-Ping

    2003-03-01

    The reliability of kinetic substrate quantification by nonlinear fitting of the enzyme reaction curve to the integrated Michaelis-Menten equation was investigated by both simulation and preliminary experimentation. For simulation, product absorptivity epsilon was 3.00 mmol(-1) L cm(-1) and K(m) was 0.10 mmol L(-1), and uniform absorbance error sigma was randomly inserted into the error-free reaction curve of product absorbance A(i) versus reaction time t(i) calculated according to the integrated Michaelis-Menten equation. The experimental reaction curve of arylesterase acting on phenyl acetate was monitored by phenol absorbance at 270 nm. Maximal product absorbance A(m) was predicted by nonlinear fitting of the reaction curve to Eq. (1) with K(m) as constant. There were unique A(m) for best fitting of both the simulated and experimental reaction curves. Neither the error in reaction origin nor the variation of enzyme activity changed the background-corrected value of A(m). But the range of data under analysis, the background absorbance, and absorbance error sigma had an effect. By simulation, A(m) from 0.150 to 3.600 was predicted with reliability and linear response to substrate concentration when there was 80% consumption of substrate at sigma of 0.001. Restriction of absorbance to 0.700 enabled A(m) up to 1.800 to be predicted at sigma of 0.001. Detection limit reached A(m) of 0.090 at sigma of 0.001. By experimentation, the reproducibility was 4.6% at substrate concentration twice the K(m), and A(m) linearly responded to phenyl acetate with consistent absorptivity for phenol, and upper limit about twice the maximum of experimental absorbance. These results supported the reliability of this new kinetic method for enzymatic analysis with enhanced upper limit and precision.

  4. Kinetics of Circulating Plasma Cell-Free DNA in Paediatric Classical Hodgkin Lymphoma

    Science.gov (United States)

    Primerano, Simona; Burnelli, Roberta; Carraro, Elisa; Pillon, Marta; Elia, Caterina; Farruggia, Piero; Sala, Alessandra; Vinti, Luciana; Buffardi, Salvatore; Basso, Giuseppe; Mascarin, Maurizio; Mussolin, Lara

    2016-01-01

    Levels of plasma cell-free DNA (cfDNA) of a large series of children with classical Hodgkin lymphoma (cHL) were evaluated and analyzed at diagnosis and during chemotherapy treatment in relation with clinical characteristics. CfDNA levels in cHL patients were significantly higher compared with controls (p=0.002). CfDNA at diagnosis was correlated with presence of B symptoms (p=0.027) and high erythrocyte sedimentation rate (p=0.049). We found that the increasing of plasma cfDNA after first chemotherapy cycle seems to be associated with a worse prognosis (p=0.049). Levels of plasma cfDNA might constitute an interesting non-invasive tool in cHL patients' management. PMID:26918050

  5. Measuring kinetic isotope effects in enzyme reactions using time-resolved electrospray mass spectrometry.

    Science.gov (United States)

    Liuni, Peter; Olkhov-Mitsel, Ekaterina; Orellana, Arturo; Wilson, Derek J

    2013-04-01

    Kinetic isotope effect (KIE) measurements are a powerful tool for studying enzyme mechanisms; they can provide insights into microscopic catalytic processes and even structural constraints for transition states. However, KIEs have not come into widespread use in enzymology, due in large part to the requirement for prohibitively cumbersome experimental procedures and daunting analytical frameworks. In this work, we introduce time-resolved electrospray ionization mass spectrometry (TRESI-MS) as a straightforward, precise, and inexpensive method for measuring KIEs. Neither radioisotopes nor large amounts of material are needed and kinetic measurements for isotopically "labeled" and "unlabeled" species are acquired simultaneously in a single "competitive" assay. The approach is demonstrated first using a relatively large isotope effect associated with yeast alcohol dehydrogenase (YADH) catalyzed oxidation of ethanol. The measured macroscopic KIE of 2.19 ± 0.05 is consistent with comparable measurements in the literature but cannot be interpreted in a way that provides insights into isotope effects in individual microscopic steps. To demonstrate the ability of TRESI-MS to directly measure intrinsic KIEs and to characterize the precision of the technique, we measure a much smaller (12)C/(13)C KIE associated specifically with presteady state acylation of chymotrypsin during hydrolysis of an ester substrate.

  6. Enzyme

    Science.gov (United States)

    Enzymes are complex proteins that cause a specific chemical change in all parts of the body. For ... use them. Blood clotting is another example of enzymes at work. Enzymes are needed for all body ...

  7. Research progress of the atypical kinetic profiles of cytochrome P450 enzymes%细胞色素P450酶的非典型动力学研究进展

    Institute of Scientific and Technical Information of China (English)

    曾彩雯; 何芳; 夏春华; 熊玉卿

    2012-01-01

    Cytochrome P450 enzymes are composed of many isozyraes and involved in the biotransformation of both exogenous and endogenous substances. A growing number of studies have found that the P450 enzymes do not always follow the classical Michaelis-Menten kinetics, but show atypical kinetic behavior, which is also the current research hotspot. In this paper, the category and mechanisms of atypical kinetics of the P450 enzyme were reviewed, providing theoretical basis for the research of enzyme kinetics.%细胞色素P450酶是一组由许多同工酶组成的超基因大家族,在外源性和内源性物质的代谢中起着极其重要的作用.越来越多的研究发现,P450酶并不总是遵循经典的米氏动力学,动力学模式经常以别构形式出现,即表现出非典型动力学行为,这也是当今的一个研究热点.本文就P450酶的非典型动力学表现形式及其发生机制做一综述,为酶动力学研究提供科学依据.

  8. Kinetic modelling of enzyme inactivation Kinetics of heat inactivation of the extracellular proteinase from Pseudomonas fluorescens 22F.

    NARCIS (Netherlands)

    Schokker, E.P.

    1997-01-01

    The kinetics of heat inactivation of the extracellular proteinase from Pseudomonas fluorescens 22F was studied. It was established, by making use of kinetic modelling, that heat inactivation in the temperature range 35 - 70 °C was most likely caused by intermolecular autoproteolysis, where unfolded

  9. Different enzyme kinetics of midazolam in recombinant CYP3A4 microsomes from human and insect sources.

    Science.gov (United States)

    Christensen, Hege; Mathiesen, Liv; Postvoll, Lillian W; Winther, Bjørn; Molden, Espen

    2009-01-01

    In vitro drug metabolism techniques with human CYP c-DNA expressed systems are frequently used to predict human drug metabolism in vivo. The aim of this study was to compare midazolam enzyme kinetics in recombinant expressed CYP3A4 microsomes from human and insect cells. The amounts of 1'- hydroxymidazolam and 4-hydroxymidazolam formed in CYP3A4 microsomes from transfected human liver epithelial cells (T5-3A4 microsomes) and baculovirus-infected insect cells (with and without coexpressed cytochrome b(5)) were analysed by LC-MS. Enzyme kinetic parameters were estimated by nonlinear regression. Mean K(m) for the formation of 1'-hydroxymidazolam was 3- and 4-fold higher in T5-3A4 microsomes than in insect microsomes (pmicrosomes was reflected by significantly lower Cl(int) compared to insect microsomes (pmicrosomes displayed Michaelis-Menten kinetics, while insect microsomes showed substrate inhibition kinetics. The different enzyme kinetics of midazolam observed in recombinant CYP3A4 microsomes from human and insect sources, especially the substantially higher K(m) obtained in human microsomes compared to insect microsomes, should be further evaluated since it may have implications for correlations to in vivo situation.

  10. Enzyme kinetics by directly imaging a porous silicon microfluidic reactor using desorption/ionization on silicon mass spectrometry.

    Science.gov (United States)

    Nichols, Kevin P; Azoz, Seyla; Gardeniers, Han J G E

    2008-11-01

    Enzyme kinetics were obtained in a porous silicon microfluidic channel by combining an enzyme and substrate droplet, allowing them to react and deposit a small amount of residue on the channel walls, and then analyzing this residue by directly ionizing the channel walls using a matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) laser source. The porous silicon of the channel walls functions in a manner analogous to the matrix in MALDI-MS, and is referred to as a desorption/ionization on silicon mass spectrometry (DIOS-MS) target when used in this configuration. Mass spectrometry signal intensity of substrate residue correlates with relative concentration, and position in the microchannel correlates with time, thus allowing determination of kinetic parameters. The system is especially suitable for initial reaction velocity determination. This microreactor is broadly applicable to time-resolved kinetic assays as long as at least one substrate or product of the reaction is ionizable by DIOS-MS.

  11. Universality of Poisson indicator and Fano factor of transport event statistics in ion channels and enzyme kinetics.

    Science.gov (United States)

    Chaudhury, Srabanti; Cao, Jianshu; Sinitsyn, Nikolai A

    2013-01-17

    We consider a generic stochastic model of ion transport through a single channel with arbitrary internal structure and kinetic rates of transitions between internal states. This model is also applicable to describe kinetics of a class of enzymes in which turnover events correspond to conversion of substrate into product by a single enzyme molecule. We show that measurement of statistics of single molecule transition time through the channel contains only restricted information about internal structure of the channel. In particular, the most accessible flux fluctuation characteristics, such as the Poisson indicator (P) and the Fano factor (F) as function of solute concentration, depend only on three parameters in addition to the parameters of the Michaelis-Menten curve that characterizes average current through the channel. Nevertheless, measurement of Poisson indicator or Fano factor for such renewal processes can discriminate reactions with multiple intermediate steps as well as provide valuable information about the internal kinetic rates.

  12. Facial-muscle weakness, speech disorders and dysphagia are common in patients with classic infantile Pompe disease treated with enzyme therapy

    NARCIS (Netherlands)

    C.M. Van Gelder (Carin); C.I. van Capelle (Carine); B.J. Ebbink; I. Moor-van Nugteren; J.M.P. van den Hout (Johanna); M.M. Hakkesteegt (Marieke); P.A. van Doorn (Pieter); I.F.M. de Coo (René); A.J.J. Reuser (Arnold); H.H.W. de Gier (Henriette); A.T. van der Ploeg (Ans)

    2012-01-01

    textabstractClassic infantile Pompe disease is an inherited generalized glycogen storage disorder caused by deficiency of lysosomal acid α-glucosidase. If left untreated, patients die before one year of age. Although enzyme-replacement therapy (ERT) has significantly prolonged lifespan, it has also

  13. Computer controlled automated assay for comprehensive studies of enzyme kinetic parameters.

    Directory of Open Access Journals (Sweden)

    Felix Bonowski

    Full Text Available Stability and biological activity of proteins is highly dependent on their physicochemical environment. The development of realistic models of biological systems necessitates quantitative information on the response to changes of external conditions like pH, salinity and concentrations of substrates and allosteric modulators. Changes in just a few variable parameters rapidly lead to large numbers of experimental conditions, which go beyond the experimental capacity of most research groups. We implemented a computer-aided experimenting framework ("robot lab assistant" that allows us to parameterize abstract, human-readable descriptions of micro-plate based experiments with variable parameters and execute them on a conventional 8 channel liquid handling robot fitted with a sensitive plate reader. A set of newly developed R-packages translates the instructions into machine commands, executes them, collects the data and processes it without user-interaction. By combining script-driven experimental planning, execution and data-analysis, our system can react to experimental outcomes autonomously, allowing outcome-based iterative experimental strategies. The framework was applied in a response-surface model based iterative optimization of buffer conditions and investigation of substrate, allosteric effector, pH and salt dependent activity profiles of pyruvate kinase (PYK. A diprotic model of enzyme kinetics was used to model the combined effects of changing pH and substrate concentrations. The 8 parameters of the model could be estimated from a single two-hour experiment using nonlinear least-squares regression. The model with the estimated parameters successfully predicted pH and PEP dependence of initial reaction rates, while the PEP concentration dependent shift of optimal pH could only be reproduced with a set of manually tweaked parameters. Differences between model-predictions and experimental observations at low pH suggest additional protonation

  14. Enzyme Kinetics: A critique of the quasi-steady-state approximation

    CERN Document Server

    Bhattacharyya, Kamal

    2013-01-01

    The standard two-step model of homogeneous-catalyzed reactions had been theoretically analyzed at various levels of approximations from time to time. The primary aim was to check the validity of the quasi-steady-state approximation, and hence emergence of the Michaelis-Menten kinetics, with various substrate-enzyme ratios. But, conclusions vary. We solve here the desired set of coupled nonlinear differential equations by invoking a new set of dimensionless variables. Approximate solutions are obtained via the power-series method aided by Pade approximants. The scheme works very successfully in furnishing the initial dynamics at least up to the region where existence of any steady state can be checked. A few conditions for its validity are put forward and tested against the findings. Temporal profiles of the substrate and the product are analyzed in addition to that of the complex to gain further insights into legitimacy of the above approximation. Some recent observations like the reactant stationary approxim...

  15. Determination of the Effects of Hazelnut Husk and Tea Waste Treatments on Urease Enzyme Activity and Its Kinetics in Soil

    OpenAIRE

    Rıdvan KIZILKAYA; EKBERLİ, İmanverdi

    2008-01-01

    In this research, the effects of 5% treatment of hazelnut husk (HH) and tea production waste (TEW) to clay loam soil on urease enzyme activity and its kinetics were determined in a 30-day soil incubation experiment. For this purpose, kinetic parameters (Vmax, Km, and Vmax/Km,) were calculated by determining urease activity in organic wastes treated soils in different substrate concentrations (0%, 1%, 2%, 4%, 6%, 8%, 10%, and 12%), incubation periods (0, 1, 2, 3, 4, 5 and 6 h), and incubation ...

  16. Enzyme kinetics and distinct modulation of the protein kinase N family of kinases by lipid activators and small molecule inhibitors

    Science.gov (United States)

    Falk, Matthew D.; Liu, Wei; Bolaños, Ben; Unsal-Kacmaz, Keziban; Klippel, Anke; Grant, Stephan; Brooun, Alexei; Timofeevski, Sergei

    2014-01-01

    The PKN (protein kinase N) family of Ser/Thr protein kinases regulates a diverse set of cellular functions, such as cell migration and cytoskeletal organization. Inhibition of tumour PKN activity has been explored as an oncology therapeutic approach, with a PKN3-targeted RNAi (RNA interference)-derived therapeutic agent in Phase I clinical trials. To better understand this important family of kinases, we performed detailed enzymatic characterization, determining the kinetic mechanism and lipid sensitivity of each PKN isoform using full-length enzymes and synthetic peptide substrate. Steady-state kinetic analysis revealed that PKN1–3 follows a sequential ordered Bi–Bi kinetic mechanism, where peptide substrate binding is preceded by ATP binding. This kinetic mechanism was confirmed by additional kinetic studies for product inhibition and affinity of small molecule inhibitors. The known lipid effector, arachidonic acid, increased the catalytic efficiency of each isoform, mainly through an increase in kcat for PKN1 and PKN2, and a decrease in peptide KM for PKN3. In addition, a number of PKN inhibitors with various degrees of isoform selectivity, including potent (Ki<10 nM) and selective PKN3 inhibitors, were identified by testing commercial libraries of small molecule kinase inhibitors. This study provides a kinetic framework and useful chemical probes for understanding PKN biology and the discovery of isoform-selective PKN-targeted inhibitors. PMID:27919031

  17. In vitro glucuronidation kinetics of deoxynivalenol by human and animal microsomes and recombinant human UGT enzymes.

    Science.gov (United States)

    Maul, Ronald; Warth, Benedikt; Schebb, Nils Helge; Krska, Rudolf; Koch, Matthias; Sulyok, Michael

    2015-06-01

    The mycotoxin deoxynivalenol (DON), formed by Fusarium species, is one of the most abundant mycotoxins contaminating food and feed worldwide. Upon ingestion, the majority of the toxin is excreted by humans and animal species as glucuronide conjugate. First in vitro data indicated that DON phase II metabolism is strongly species dependent. However, kinetic data on the in vitro metabolism as well as investigations on the specific enzymes responsible for DON glucuronidation in human are lacking. In the present study, the DON metabolism was investigated using human microsomal fractions and uridine-diphosphoglucuronyltransferases (UGTs) as well as liver microsomes from five animal species. Only two of the twelve tested human recombinant UGTs led to the formation of DON glucuronides with a different regiospecificity. UGT2B4 predominantly catalyzed the formation of DON-15-O-glucuronide (DON-15GlcA), while for UGT2B7 the DON-3-O-glucuronide (DON-3GlcA) metabolite prevailed. For human UGTs, liver, and intestinal microsomes, the glucuronidation activities were low. The estimated apparent intrinsic clearance (Clapp,int) for all human UGT as well as tissue homogenates was microsomes, moderate Clapp,int between 1.5 and 10 mL/min mg protein were calculated for carp, trout, and porcine liver. An elevated glucuronidation activity was detected for rat and bovine liver microsomes leading to Clapp,int between 20 and 80 mL/min mg protein. The obtained in vitro data points out that none of the animal models is suitable for estimating the human DON metabolism with respect to the metabolite pattern and formation rate.

  18. The identification of enzyme targets for the optimization of a valine producing Corynebacterium glutamicum strain using a kinetic model.

    Science.gov (United States)

    Magnus, Jørgen Barsett; Oldiges, Marco; Takors, Ralf

    2009-01-01

    The enzyme targets for the rational optimization of a Corynebacterium glutamicum strain constructed for valine production are identified by analyzing the control of flux in the valine/leucine pathway. The control analysis is based on measurements of the intracellular metabolite concentrations and on a kinetic model of the reactions in the investigated pathway. Data-driven and model-based methods are used and evaluated against each other. The approach taken gives a quantitative evaluation of the flux control and it is demonstrated how the understanding of flux control is used to reach specific recommendations for strain optimization. The flux control coefficients (FCCs) with respect to the valine excretion rate were calculated, and it was found that the control is distributed mainly between the acetohydroxyacid synthase enzyme (FCC = 0.32), the branched chain amino acid transaminase (FCC = 0.27), and the exporting translocase (FCC = 0.43). The availability of the precursor pyruvate has substantial influence on the valine flux, whereas the cometabolites are less important as demonstrated by the calculation of the respective response coefficients. The model is further used to make in-silico predictions of the change in valine flux following a change in enzyme level. A doubling of the enzyme level of valine translocase will result in an increase in valine flux of 31%. By optimizing the enzyme levels with respect to valine flux it was found that the valine flux can be increased by a factor 2.5 when the optimal enzyme levels are implemented.

  19. Catalytic efficiency of dehaloperoxidase A is controlled by electrostatics--application of the vibrational Stark effect to understand enzyme kinetics.

    Science.gov (United States)

    Schkolnik, Gal; Utesch, Tillmann; Zhao, Junjie; Jiang, Shu; Thompson, Matthew K; Mroginski, Maria-Andrea; Hildebrandt, Peter; Franzen, Stefan

    2013-01-18

    The vibrational Stark effect is gaining popularity as a method for probing electric fields in proteins. In this work, we employ it to explain the effect of single charge mutations in dehaloperoxidase-hemoglobin A (DHP A) on the kinetics of the enzyme. In a previous communication published in this journal (BBRC 2012, 420, 733-737) it has been shown that an increase in the overall negative charge of DHP A through mutation causes a decrease in its catalytic efficiency. Here, by labeling the protein with 4-mercaptobenzonitrile (MBN), a Stark probe molecule, we provide further evidence that the diffusion control of the catalytic process arises from the electrostatic repulsion between the enzyme and the negatively charged substrate. The linear correlation observed between the nitrile stretching frequency of the protein-bound MBN and the catalytic efficiency of the single-site mutants of the enzyme indicates that electrostatic interactions play a dominant role in determining the catalytic efficiency of DHP A.

  20. A New Enzyme Immunoassay for the Quantitative Determination of Classical Autotaxins (ATXα, ATXβ, and ATXγ and Novel Autotaxins (ATXδ and ATXε.

    Directory of Open Access Journals (Sweden)

    Yasunori Tokuhara

    Full Text Available Autotaxin (ATX is a secreted enzyme that converts lysophosphatidylcholine to lysophosphatidic acid, a potent bioactive lipid mediator, through its lysophospholipase D activity. Although five alternative splicing isoforms of ATX have been identified as ATXα, ATXβ, ATXγ, ATXδ, and ATXε and the expression patterns of each isoform differ among several tissues, the clinical significance of each isoform remains to be elucidated.Anti-ATXβ and anti-ATXδ monoclonal antibodies were produced by immunization with recombinant human ATXβ and ATXδ expressed using a baculovirus system, respectively. We then developed enzyme immunoassays to measure the serum concentrations of "classical ATX" (ATXα, ATXβ, and ATXγ and "novel ATX" (ATXδ and ATXε antigens and evaluated the usefulness of these assays using human serum samples.The with-run and between-run precision, interference, detection limit, and linearity studies for the present assay were well validated. In healthy subjects, the serum concentrations of classical ATX and novel ATX were significantly (P < 0.01 higher in women than in men, while the ratios of classical ATX or novel ATX to total ATX were not different between women and men. The concentrations of both classical ATX and novel ATX in normal pregnant subjects and patients with chronic liver diseases or follicular lymphoma were significantly higher than those in healthy subjects, while the ratio of both ATX isoforms to total ATX did not vary among these groups.We have developed a new enzyme immunoassay to determine the concentrations of classical ATX and novel ATX in human serum. These assays may be helpful for elucidating the distinct functional roles of each ATX isoform, which are largely unknown at present.

  1. Sugarcane bagasse pretreatment using three imidazolium-based ionic liquids; mass balances and enzyme kinetics

    Directory of Open Access Journals (Sweden)

    Karatzos Sergios

    2012-08-01

    Full Text Available Abstract Background Effective pretreatment is key to achieving high enzymatic saccharification efficiency in processing lignocellulosic biomass to fermentable sugars, biofuels and value-added products. Ionic liquids (ILs, still relatively new class of solvents, are attractive for biomass pretreatment because some demonstrate the rare ability to dissolve all components of lignocellulosic biomass including highly ordered (crystalline cellulose. In the present study, three ILs, 1-butyl-3-methylimidazolium chloride ([C4mim]Cl, 1-ethyl-3-methylimidazolium chloride ([C2mim]Cl, 1-ethyl-3-methylimidazolium acetate ([C2mim]OAc are used to dissolve/pretreat and fractionate sugarcane bagasse. In these IL-based pretreatments the biomass is completely or partially dissolved in ILs at temperatures greater than 130°C and then precipitated by the addition of an antisolvent to the IL biomass mixture. For the first time mass balances of IL-based pretreatments are reported. Such mass balances, along with kinetics data, can be used in process modelling and design. Results Lignin removals of 10% mass of lignin in bagasse with [C4mim]Cl, 50% mass with [C2mim]Cl and 60% mass with [C2mim]OAc, are achieved by limiting the amount of water added as antisolvent to 0.5 water:IL mass ratio thus minimising lignin precipitation. Enzyme saccharification (24 h, 15FPU yields (% cellulose mass in starting bagasse from the recovered solids rank as: [C2mim]OAc(83% > >[C2mim]Cl(53% = [C4mim]Cl(53%. Composition of [C2mim]OAc-treated solids such as low lignin, low acetyl group content and preservation of arabinosyl groups are characteristic of aqueous alkali pretreatments while those of chloride IL-treated solids resemble aqueous acid pretreatments. All ILs are fully recovered after use (100% mass as determined by ion chromatography. Conclusions In all three ILs regulated addition of water as an antisolvent effected a polysaccharide enriched precipitate since some of the lignin

  2. Structure of enzyme-bound substrates: resonance Raman and kinetic evidence for differential enzyme-substrate contacts in N-(Pentafluoro-benzoyl)glycine dithioacyl and thioacyl papain

    Science.gov (United States)

    Lee, H.; Angus, R. H.; Storer, A. C.; Carey, P. R.

    1989-12-01

    Resonance Raman (RR) spectroscopy is used to probe the structure of the substrate in the substrate-enzyme complex N-pentafluorobenzoyl) glycine (dithioacyl) papain (C 6F 5C(=O) NHCH 2C(=S)S-papain). This system was chosen since the high electron withdrawing capacity of the C 6F 5 group markedly affects electron density of the -NH- moiety which, in turn, is known to change catalytic activity. The RR spectrum of the enzyme-substrate complex is interpreted by reference to the model compound N-(pentafluorobenzoyl) glycine ethyl dithioester (C 6F 5(CO))NHCH 2C(=S)SC 2H 5. The RR spectra of this compound in aqueous or organic solvents can be understood in terms of the known conformational states of N-acylglycine dithioesters. Comparison of model with enzyme-substrate RR spectra shows that the substrate is binding in the active site in a conformer known as conformer B characterized by a small-NHCH 2CS(thiol) torsional angle and close N-to-S (thiol) contact. Kinetic rate-structure correlations are developed involving k3, the rate constant for deacylation, and the strength of the N-to-S (thiol) interaction. N-(Pentafluorobenzoyl) glycine dithioacyl papain fits the rate-structure correlation whereas the corresponding pentafluorobenzoyl glycine thiol intermediate does not. It is proposed that the difference in the size of the CS compared to the CO group brings about a small change in the dithioacyl papain compared to the thiolacyl papain conformation such that enzyme-substrate contacts involving ortho and meta F atoms in the thiol acyl enzyme case are weakened or removed in the case of the dithioacyl papain.

  3. Kinetic and structural analysis of enzyme sliding on a substrate: multiple attack in beta-amylase.

    Science.gov (United States)

    Ishikawa, Kazuhiko; Nakatani, Hiroshi; Katsuya, Yoshio; Fukazawa, Chikafusa

    2007-01-23

    Beta-amylase (EC 3.2.1.2) is starch-hydrolyzing exo-type enzyme that can catalyze the successive liberation of beta-maltose from the nonreducing ends of alpha-1,4-linked glucopyranosyl polymers. There is a well-known phenomenon called multiple or repetitive attack where the enzyme releases several maltose molecules in a single enzyme-substrate complex. In order to understand it further, we examined the beta-amylase-catalyzed reaction using maltooligosaccharides. The Monte Carlo method was applied for simulation of the beta-amylase-catalyzed reaction including the multiple attack mechanism. Through site-directed mutagenesis, we have successfully prepared a mutant enzyme which may be simulated as a multiple attack action reduced one with retaining significant hydrolytic activity. From the results of X-ray structure analysis of the mutant enzyme, it was clarified that one carboxyl residue plays a very important role in the multiple attack. The multiple attack action needs the force of enzyme sliding on the substrate. In addition, it is important for the multiple attack that the enzyme and substrate have the characteristics of a stable productive substrate-enzyme complex through a hydrogen bond between the nonreducing end of the substrate and the carboxyl residue of the enzyme.

  4. Nonlinear temperature sensitivity of enzyme kinetics explains canceling effect—a case study on loamy haplic Luvisol

    Science.gov (United States)

    Razavi, Bahar S.; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2015-01-01

    The temperature sensitivity of enzymes responsible for organic matter decomposition in soil is crucial for predicting the effects of global warming on the carbon cycle and sequestration. We tested the hypothesis that differences in temperature sensitivity of enzyme kinetic parameters Vmax and Km will lead to a canceling effect: strong reduction of temperature response of catalytic reactions. Short-term temperature response of Vmax and Km of three hydrolytic enzymes responsible for decomposition of cellulose (β-glucosidase, cellobiohydrolase) and hemicelluloses (xylanase) were analyzed in situ from 0 to 40°C. The apparent activation energy varied between enzymes from 20.7 to 35.2 kJ mol−1 corresponding to the Q10 values of the enzyme activities of 1.4–1.9 (with Vmax-Q10 1.0–2.5 and Km-Q10 0.94–2.3). Temperature response of all tested enzymes fitted well to the Arrhenius equation. Despite that, the fitting of Arrhenius model revealed the non-linear increase of two cellulolytic enzymes activities with two distinct thresholds at 10–15°C and 25–30°C, which were less pronounced for xylanase. The nonlinearity between 10 and 15°C was explained by 30–80% increase in Vmax. At 25–30°C, however, the abrupt decrease of enzyme-substrate affinity was responsible for non-linear increase of enzyme activities. Our study is the first demonstrating nonlinear response of Vmax and Km to temperature causing canceling effect, which was most strongly pronounced at low substrate concentrations and at temperatures above 15°C. Under cold climate, however, the regulation of hydrolytic activity by canceling in response to warming is negligible because canceling was never observed below 10°C. The canceling, therefore, can be considered as natural mechanism reducing the effects of global warming on decomposition of soil organics at moderate temperatures. The non-linearity of enzyme responses to warming and the respective thresholds should therefore be investigated for

  5. Estimating the turnover number in enzyme kinetic reactions using transient and stationary state data

    Directory of Open Access Journals (Sweden)

    Sibel Uludag-Demirer

    2009-12-01

    Full Text Available Substrate and product concentration data obtained by simulating enzyme-substrate reaction rate equations were used to test two proposed kinetic rate constant estimation techniques in this study. In the first technique, the turnover number, k3, was calculated using early transient time domain data, which are difficult to obtain experimentally. The technique used an iterative approach to calculate k3 with a pair of data and the value of k3 could be retrieved with 35% error. The second technique calculated k3 using stationary domain data and the value of k3 could be retrieved with less than 5% error. This second technique also offered internal consistency in the calculation of k3 by calculating k3 both from the intercept and the slope of the linear plot derived in this study. A series of sensitivity analyses was conducted to understand the robustness of the second technique in estimating k3 from simulated data to the changes in the reaction rate constants (k1, k2, and k3 and the initial concentration of enzyme used for simulation. It was found that the second technique generally worked well in the estimation of k3 except for the simulated data for fast substrate conversions such as in the large k3 and [E]0 cases . This latter method, thus, shows promise for the use of late time experimental substrate/product concentration data to obtain k3. Exclusively using late time data avoids the need for difficult and expensive rapid early time measurement techniques for estimating k3. Once a reasonable estimate for k3 is obtained, the initial enzyme value can easily be determined from the maximum velocity constant established from fitting the Michaelis-Menten or Briggs-Haldane equations to substrate and product stationary state domain (late time data. While the first technique can estimate k3 with only one point in the transient domain, it is suggested that the second method generally be favored since it only requires late-time stationary domain data and

  6. Kinetic characteristics of polygalacturonase enzymes hydrolyzing galacturonic acid oligomers using isothermal titration calorimetry

    Science.gov (United States)

    Polygalacturonase enzymes hydrolyze the polygalacturonic acid chains found in pectin. Interest in polygalacturonase enzymes continues as they are useful in a number of industrial processes and conversely, detrimental, as they are involved in maceration of economically important crops. While a good...

  7. Escherichia coli Phosphoenolpyruvate-Dependent Phosphotransferase System : Equilibrium Kinetics and Mechanism of Enzyme I Phosphorylation

    NARCIS (Netherlands)

    Hoving, H; Lolkema, Juke S.; Robillard, George T.

    1981-01-01

    The phosphorylation of enzyme I from the Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system was studied by means of isotope exchange between phosphoenolpyruvate and pyruvate. Experiments monitoring 1H-2H exchange showed that enzyme I phosphorylation is accompanied by the transf

  8. Degradation kinetics of forchlorfenuron in typical grapevine soils of India and its influence on specific soil enzyme activities.

    Science.gov (United States)

    Banerjee, Kaushik; Dasgupta, Soma; Oulkar, Dasharath P; Patil, Sangram H; Adsule, Pandurang G

    2008-05-01

    The rate of degradation of forchlorfenuron, a cytokinin-based plant growth regulator (PGR) was explored in typical grapevine soils of India with simultaneous evaluation of its effect on biochemical attributes of the test soils in terms of the activities of specific soil microbial enzymes. In all the test soils, namely clay, sandy-loam and silty-clay, the dissipation rate was faster at the beginning, which slowed down with time, indicating a non-linear pattern of degradation. Degradation in soils could best be explained by two-compartment 1st+1st order kinetics with half-life ranging between 4-10 days. The results suggest that organic matter might be playing a major role in influencing the rate of degradation of forchlorfenuron in soil. The rate of degradation in sandy-loam soil was fastest followed by clay and silty-clay soils, respectively. Comparison of the rate of degradation in natural against sterilized soils suggests that microbial degradation might be the major pathway of residue dissipation. Changes in soil enzyme activities as a consequence of forchlorfenuron treatment were studied for extra-cellular enzymes namely acid phosphatase, alkaline phosphatase and beta -glucosidase and intracellular enzyme-dehydrogenase. Although small changes in enzyme activities were observed, forchlorfenuron did not have any significant deleterious effect on the enzymatic activity of the test soils. Simple correlation studies between degradation percentage and individual enzyme activities did not establish any significant relationships. The pattern and change of enzyme activity was primarily the effect of the incubation period rather than the effect of forchlorfenuron itself.

  9. Studies on NADPH-cytochrome c reductase. II. Steady-state kinetic properties of the crystalline enzyme from ale yeast.

    Science.gov (United States)

    Tryon, E; Kuby, S A

    1984-01-01

    From a study of the steady-state kinetics (at pH 7.6, 30 degrees C) of the reduction of cytochrome c, a 'ping-pong' mechanism may be postulated for the crystalline NADPH-cytochrome c reductase from ale yeast, Saccharomyces cerevisiae [1], a result derivable from a three-substrate ordered system with a rapid equilibrium random sequence in substrates, NADPH and FAD, followed by reactions of the third substrate, Cyt C3+. On this basis, estimates for the kinetic parameters were made together with the inhibitor dissociation constants for NADP+ (competitive with respect to NADPH as variable substrate, but noncompetitive with respect to cytochrome c3+ as the variable substrate). A noncompetitive type of inhibition was also found for cytochrome c2+ with NADPH as variable substrate, in confirmation of the proposed mechanism. With 2,6-dichloroindophenol as the acceptor, in place of cytochrome c3+, a value for KNADPH could be estimated which agreed with that estimated above, with cytochrome c3+ as the acceptor, again, in confirmation of the postulated mechanism. The reactions with molecular O2 catalyzed by the enzyme with NADPH as the reductant have been studied polarographically, and its Km for O2 estimated to be about 0.15 mmol/l at pH 7.6, 25 degrees C. The product of the reaction appears to be H2O2, which acts as a noncompetitive inhibitor for NADPH (Ki = 0.5 mmol/l), and tentatively an enzyme ternary complex containing oxygen and FADoh (semiquinone of FAD) may be assumed to be the kinetically important intermediate, which may be postulated to be in quasi-equilibrium with an enzyme ternary complex containing Oo2 (superoxide) and FAD.

  10. Approaching a Conceptual Understanding of Enzyme Kinetics and Inhibition: Development of an Active Learning Inquiry Activity for Prehealth and Nonscience Majors

    Science.gov (United States)

    House, Chloe; Meades, Glen; Linenberger, Kimberly J.

    2016-01-01

    Presented is a guided inquiry activity designed to be conducted with prenursing students using an analogous system to help develop a conceptual understanding of factors impacting enzyme kinetics and the various types of enzyme inhibition. Pre- and postconceptual understanding evaluations and effectiveness of implementation surveys were given to…

  11. Application of A Microfluidic Tool for the Determination of Enzyme Kinetics

    OpenAIRE

    Ringborg, Rolf H.; Woodley, John M; Gernaey, Krist; Krühne, Ulrich

    2015-01-01

    Med biokatalyse er det muligt at syntetisere og producere værdifulde kemikalier under mildebetingelser. Ved udvikling af nye processer, bliver enzymer modificeret til at katalysere ønskede produkter med stor succes. Nu om dage introduceres mutationer i enzymer, derved dannes mange nye mutanter og der søges herefter iblandt disse. Med high throughput screening kan man udføre screening af millioner af mutanter pr. dag. Der er derfor høj sandsynlighed for at finde enzym mutanter med egnede egens...

  12. Numerical stability of finite difference algorithms for electrochemical kinetic simulations: Matrix stability analysis of the classic explicit, fully implicit and Crank-Nicolson methods and typical problems involving mixed boundary conditions

    DEFF Research Database (Denmark)

    Bieniasz, Leslaw K.; Østerby, Ole; Britz, Dieter

    1995-01-01

    The stepwise numerical stability of the classic explicit, fully implicit and Crank-Nicolson finite difference discretizations of example diffusional initial boundary value problems from electrochemical kinetics has been investigated using the matrix method of stability analysis. Special attention...

  13. Kinetics of enzyme-catalyzed cross-linking of feruloylated arabinan from sugar beet

    DEFF Research Database (Denmark)

    Abang Zaidel, Dayang Norulfairuz; Arnous, Anis; Holck, Jesper

    2011-01-01

    the kinetics of HRP catalyzed cross-linking of FA esterified to α-(1,5)-linked arabinans are affected by the length of the arabinan chains carrying the feruloyl substitutions. The kinetics of the HRP-catalyzed cross-linking of four sets of arabinan samples from sugar beet pulp, having different molecular...... weights and hence different degrees of polymerization, were monitored by the disappearance of FA absorbance at 316 nm. MALDI-TOF/TOF-MS analysis confirmed that the sugar beet arabinans were feruloyl-substituted, and HPLC analysis verified that the amounts of diFAs increased when FA levels decreased...

  14. On the dynamics of immobilized enzyme kinetics in a microreactor: A study of AP-catalyzed reactions

    Directory of Open Access Journals (Sweden)

    Pratap R Patnaik

    2011-11-01

    Full Text Available Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin-top:0in; mso-para-margin-right:0in; mso-para-margin-bottom:10.0pt; mso-para-margin-left:0in; line-height:115%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;} The kinetics of immobilized enzyme-catalyzed reactions in microreactors differ from those in macro-scale reactors. Recognizing this, a recent study (Patnaik 2011 based on a new interpretation of the kinetics of AP-catalyzed reactions showed that dynamic behavior is feasible only certain loci relating key kinetic parameters. That work has been extended here, and the kinetic parameters have now been related to bulk phase concentrations, thereby providing a link with the reaction system per se. It has also been shown that under certain conditions the reaction may become self-quenching but either monotonically or as damped oscillations. These two studies thus establish the importance of understanding kinetic dynamics in microreactors and in selecting feasible operating conditions.

  15. The Application of Transient-State Kinetic Isotope Effects to the Resolution of Mechanisms of Enzyme-Catalyzed Reactions

    Directory of Open Access Journals (Sweden)

    Harvey F. Fisher

    2013-07-01

    Full Text Available Much of our understanding of the mechanisms of enzyme-catalyzed reactions is based on steady-state kinetic studies. Experimentally, this approach depends solely on the measurement of rates of free product appearance (d[P]/dt, a mechanistically and mathematically complex entity. Despite the ambiguity of this observed parameter, the method’s success is due in part to the elaborate rigorously derived algebraic theory on which it is based. Transient-state kinetics, on the other hand, despite its ability to observe the formation of intermediate steps in real time, has contributed relatively little to the subject due in, some measure, to the lack of such a solid mathematical basis. Here we discuss the current state of existing transient-state theory and the difficulties in its realistic application to experimental data. We describe a basic analytic theory of transient-state kinetic isotope effects in the form of three novel fundamental rules. These rules are adequate to define an extended mechanism, locating the isotope-sensitive step and identifying missing steps from experimental data. We demonstrate the application of these rules to resolved component time courses of the phenylalanine dehydrogenase reaction, extending the previously known reaction by one new prehydride transfer step and two new post hydride transfer steps. We conclude with an assessment of future directions in this area.

  16. Protein homeostasis disorders of key enzymes of amino acids metabolism: mutation-induced protein kinetic destabilization and new therapeutic strategies.

    Science.gov (United States)

    Pey, Angel L

    2013-12-01

    Many inborn errors of amino acids metabolism are caused by single point mutations affecting the ability of proteins to fold properly (i.e., protein homeostasis), thus leading to enzyme loss-of-function. Mutations may affect protein homeostasis by altering intrinsic physical properties of the polypeptide (folding thermodynamics, and rates of folding/unfolding/misfolding) as well as the interaction of partially folded states with elements of the protein homeostasis network (such as molecular chaperones and proteolytic machineries). Understanding these mutational effects on protein homeostasis is required to develop new therapeutic strategies aimed to target specific features of the mutant polypeptide. Here, I review recent work in three different diseases of protein homeostasis associated to inborn errors of amino acids metabolism: phenylketonuria, inherited homocystinuria and primary hyperoxaluria type I. These three different genetic disorders involve proteins operating in different cell organelles and displaying different structural complexities. Mutations often decrease protein kinetic stability of the native state (i.e., its half-life for irreversible denaturation), which can be studied using simple kinetic models amenable to biophysical and biochemical characterization. Natural ligands and pharmacological chaperones are shown to stabilize mutant enzymes, thus supporting their therapeutic application to overcome protein kinetic destabilization. The role of molecular chaperones in protein folding and misfolding is also discussed as well as their potential pharmacological modulation as promising new therapeutic approaches. Since current available treatments for these diseases are either burdening or only successful in a fraction of patients, alternative treatments must be considered covering studies from protein structure and biophysics to studies in animal models and patients.

  17. Distinct Enzyme-Substrate Interactions Revealed by Two Dimensional Kinetic Comparison between Dehaloperoxidase-Hemoglobin and Horseradish Peroxidase.

    Science.gov (United States)

    Zhao, Jing; Lu, Chang; Franzen, Stefan

    2015-10-08

    The time-resolved kinetics of substrate oxidation and cosubstrate H2O2 reduction by dehaloperoxidase-hemoglobin (DHP) on a seconds-to-minutes time scale was analyzed for peroxidase substrates 2,4,6-tribromophenol (2,4,6-TBP), 2,4,6-trichlorophenol (2,4,6-TCP), and ABTS. Substrates 2,4,6-TBP and 2,4,6-TCP show substrate inhibition at high concentration due to the internal binding at the distal pocket of DHP, whereas ABTS does not show substrate inhibition at any concentration. The data are consistent with an external binding site for the substrates with an internal substrate inhibitor binding site for 2,4,6-TBP and 2,4,6-TCP. We have also compared the kinetic behavior of horseradish peroxidase (HRP) in terms of kcat, Km(AH2) and Km(H2O2) using the same kinetic scheme. Unlike DHP, HRP does not exhibit any measurable substrate inhibition, consistent with substrate binding at the edge of heme near the protein surface at all substrate concentrations. The binding of substrates and their interactions with the heme iron were further compared between DHP and HRP using a competitive fluoride binding experiment, which provides a method for quantitative measurement of internal association constants associated with substrate inhibition. These experiments show the regulatory role of an internal substrate binding site in DHP from both a kinetic and competitive ligand binding perspective. The interaction of DHP with substrates as a result of internal binding actually stabilizes that protein and permits DHP to function under conditions that denature HRP. As a consequence, DHP is a tortoise, a slow but steady enzyme that wins the evolutionary race against the HRP-type of peroxidase, which is a hare, initially rapid, but flawed for this application because of the protein denaturation under the conditions of the experiment.

  18. A multiscale variational approach to the kinetics of viscous classical liquids: The coarse-grained mean field approximation

    Science.gov (United States)

    Sereda, Yuriy V.; Ortoleva, Peter J.

    2014-04-01

    A closed kinetic equation for the single-particle density of a viscous simple liquid is derived using a variational method for the Liouville equation and a coarse-grained mean-field (CGMF) ansatz. The CGMF ansatz is based on the notion that during the characteristic time of deformation a given particle interacts with many others so that it experiences an average interaction. A trial function for the N-particle probability density is constructed using a multiscale perturbation method and the CGMF ansatz is applied to it. The multiscale perturbation scheme is based on the ratio of the average nearest-neighbor atom distance to the total size of the assembly. A constraint on the initial condition is discovered which guarantees that the kinetic equation is mass-conserving and closed in the single-particle density. The kinetic equation has much of the character of the Vlasov equation except that true viscous, and not Landau, damping is accounted for. The theory captures condensation kinetics and takes much of the character of the Gross-Pitaevskii equation in the weak-gradient short-range force limit.

  19. Characterization and kinetic parameters of ethylene-forming enzyme from avocado fruit.

    Science.gov (United States)

    McGarvey, D J; Christoffersen, R E

    1992-03-25

    Biosynthesis of the phytohormone ethylene in higher plants proceeds via the following pathway: S-adenosylmethionine----1-aminocyclopropane-1-carboxylic acid (ACC)----ethylene. Ethylene-forming enzyme (EFE), the enzyme responsible for the oxidation of ACC to ethylene, has been only partially characterized in vitro. We have obtained authentic EFE activity in vitro from extracts of avocado fruit (Persea americana Mill. cv Hass). Ammonium sulfate fractionation revealed the presence of two EFE activities, which we designate as EFE1 and EFE2. EFE1 activity utilizes ACC and O2 as substrates and requires Fe(II) and ascorbate as cofactors. The enzyme has a relatively low Km (32 microM) for ACC, discriminates diastereomers of 1-amino-2-ethyl-cyclopropane-1-carboxylic acid, and is inhibited competitively by 2-aminoisobutyric acid, thus confirming its identity with authentic EFE. Activity is retained in a 100,000 x g supernatant and has a pH optimum of 7.5-8.0, suggesting a cytosolic localization.

  20. Fundamental study of the mechanism and kinetics of cellulose hydrolysis by acids and enzymes. Final report, June 1, 1978-January 31, 1981

    Energy Technology Data Exchange (ETDEWEB)

    Gong, C.S.; Chang, M.

    1981-02-01

    There are three basic enzymes (e.g., endoglucanase (C/sub x/), exoglucanase (C/sub 1/) and cellobiase) comprising the majority of extracellular cellulase enzymes produced by the cellulolytic mycelial fungi, Trichoderma reesei, and other cellulolytic microorganisms. The enzymes exhibited different mode of actions in respect to the hydrolysis of cellulose and cellulose derived oligosaccharides. In combination, these enzymes complimented each other to hydrolyze cellulose to its basic constituent, glucose. The kinetics of cellobiase were developed on the basis of applying the pseudo-steady state assumption to hydrolyze cellobiose to glucose. The results indicated that cellobiase was subjected to end-product inhibition by glucose. The kinetic modeling of exoglucanase (C/sub 1/) with respect to cellodextrins was studied. Both glucose and cellobiose were found to be inhibitors of this enzyme with cellobiose being a stronger inhibitor than glucose. Similarly, endoglucanase (C/sub x/) is subject to end-product inhibition by glucose. Crystallinity of the cellulose affects the rate of hydrolysis by cellulases. Hence, the changes in crystallinity of cellulose in relation to chemical pretreatment and enzyme hydrolysis was compared. The study of cellulase biosynthesis resulted in the conclusion that exo- and endo-glucanases are co-induced while cellobiase is synthesized independent of the other two enzymes. The multiplicity of cellulase enzymes are the end results of post-translational modification during and/or after the secretion of enzymes into growth environment.

  1. Digital simulation of scanning electrochemical microscopy approach curves to enzyme films with Michaelis-Menten kinetics.

    Science.gov (United States)

    Burchardt, Malte; Träuble, Markus; Wittstock, Gunther

    2009-06-15

    The formalism for simulating scanning electrochemical microscopy (SECM) experiments by boundary element methods in three space coordinates has been extended to allow consideration of nonlinear boundary conditions. This is achieved by iteratively refining the boundary conditions that are encoded in a boundary condition matrix. As an example, the simulations are compared to experimental approach curves in the SECM feedback mode toward samples modified with glucose oxidase (GOx). The GOx layer was prepared by the layer-by-layer assembly of polyelectrolytes using glucose oxidase as one of the polyelectrolytes. The comparison of the simulated and experimental curves showed that under a wide range of experimentally accessible conditions approximations of the kinetics at the sample by first order models yield misleading results. The approach curves differ also qualitatively from curves calculated with first order models. As a consequence, this may lead to severe deviations when such curves are fitted to first order kinetic models. The use of linear approximations to describe the enzymatic reaction in SECM feedback experiments is justified only if the ratio of the mediator and Michaelis-Menten constant is equal to or smaller than 0.1 (deviation less than 10%).

  2. Kinetics of enzyme-catalyzed hydrolysis of steam-exploded sugarcane bagasse.

    Science.gov (United States)

    Aguiar, Rodrigo Souza; Silveira, Marcos Henrique Luciano; Pitarelo, Ana Paula; Corazza, Marcos Lucio; Ramos, Luiz Pereira

    2013-11-01

    This work presents the experimental kinetic data and the fractal modeling of sugarcane bagasse steam treatment and enzymatic hydrolysis. Sugarcane bagasse (50 wt% moisture) was pretreated by autohydrolysis at 210 °C for 4 min. Acid catalysis involved the use of 9.5mg g(-1) of H2SO4 or H3PO4 in relation to the substrate dry mass at these same pretreatment conditions. Unwashed, water-washed and alkali-washed substrates were hydrolyzed at 2.0 wt% using 8 and 15 FPU g(-1) (108.22 and 199.54 mg/g) total solids of a Celluclast 1.5 L and Novozym 188 mixture (Novozymes). The fractal kinetic modeling was used to describe the effect of pretreatment and both washing processes on substrate accessibility. Water and/or alkali washing was not strictly necessary to achieve high hydrolysis efficiencies. Also, the fractal model coefficients revealed that H3PO4 was a better pretreatment catalyst under the experimental conditions used in this study, resulting in the most susceptible substrates for enzymatic hydrolysis.

  3. Beyond HTL: The Classical Kinetic Theory of Landau Damping for Selfinteracting Scalar Fields in the Broken Phase

    OpenAIRE

    1999-01-01

    The effective theory of low frequency fluctuations of selfinteracting scalar fields is constructed in the broken symmetry phase. The theory resulting from integrating fluctuations with frequencies much above the spontanously generated mass scale $(p_0>>M)$ is found to be local. Non-local dynamics, especially Landau damping emerges under the effect of fluctuations in the $p_0 \\sim M$ region. A kinetic theory of relativistic scalar gas particles interacting via their locally variable mass with ...

  4. Utilization of integrated Michaelis-Menten equations for enzyme inhibition diagnosis and determination of kinetic constants using Solver supplement of Microsoft Office Excel.

    Science.gov (United States)

    Bezerra, Rui M F; Fraga, Irene; Dias, Albino A

    2013-01-01

    Enzyme kinetic parameters are usually determined from initial rates nevertheless, laboratory instruments only measure substrate or product concentration versus reaction time (progress curves). To overcome this problem we present a methodology which uses integrated models based on Michaelis-Menten equation. The most severe practical limitation of progress curve analysis occurs when the enzyme shows a loss of activity under the chosen assay conditions. To avoid this problem it is possible to work with the same experimental points utilized for initial rates determination. This methodology is illustrated by the use of integrated kinetic equations with the well-known reaction catalyzed by alkaline phosphatase enzyme. In this work nonlinear regression was performed with the Solver supplement (Microsoft Office Excel). It is easy to work with and track graphically the convergence of SSE (sum of square errors). The diagnosis of enzyme inhibition was performed according to Akaike information criterion.

  5. Re-evaluating the kinetics of ATP hydrolysis during initiation of DNA sliding by Type III restriction enzymes.

    Science.gov (United States)

    Tóth, Júlia; Bollins, Jack; Szczelkun, Mark D

    2015-12-15

    DNA cleavage by the Type III restriction enzymes requires long-range protein communication between recognition sites facilitated by thermally-driven 1D diffusion. This 'DNA sliding' is initiated by hydrolysis of multiple ATPs catalysed by a helicase-like domain. Two distinct ATPase phases were observed using short oligoduplex substrates; the rapid consumption of ∼10 ATPs coupled to a protein conformation switch followed by a slower phase, the duration of which was dictated by the rate of dissociation from the recognition site. Here, we show that the second ATPase phase is both variable and only observable when DNA ends are proximal to the recognition site. On DNA with sites more distant from the ends, a single ATPase phase coupled to the conformation switch was observed and subsequent site dissociation required little or no further ATP hydrolysis. The overall DNA dissociation kinetics (encompassing site release, DNA sliding and escape via a DNA end) were not influenced by the second phase. Although the data simplifies the ATP hydrolysis scheme for Type III restriction enzymes, questions remain as to why multiple ATPs are hydrolysed to prepare for DNA sliding.

  6. Enzyme kinetic study of a new cardioprotective agent, KR-32570 using human liver microsomes and recombinant CYP isoforms.

    Science.gov (United States)

    Kim, Hyojin; Seo, Kyung-Ah; Kim, Hyunmi; Lee, Hye Suk; Lee, Choong-Hwan; Shin, Jae-Gook; Liu, Kwang-Hyeon

    2007-04-01

    KR-32570 (5-(2-Methoxy-5-chlorophenyl)furan-2-ylcarbonyl)guanidine) is a new cardioprotective agent for preventing ischemia-reperfusion injury. Human liver microsomal incubation of KR-32570 in the presence of NADPH resulted in the formation of two metabolites, hydroxy-KR-32570 and O-desmethyl-KR-32570. In this study, a kinetic analysis of the metabolism of two metabolites from KR-32570 was performed in human liver microsomes, and recombinant CYP1A2, and CYP3A4. The metabolism for hydroxy- and O-desmethyl-KR-32570 formation from KR-32570 by human liver microsomes was best described by a Michaelis-Menten equation and a Hill equation, respectively. The Cl(int) values of hydroxy- and O-desmethyl-KR-32570 formation were similar to each other (0.03 vs 0.04 microL/min/pmol CYP, respectively). CYP3A4 mediated the formation of hydroxy-KR-32570 from KR-32570 with Cl(int) = 0.24 microL/min/pmol CYP3A4. The intrinsic clearance for O-desmethyl-KR-32570 formation by CYP1A2 was 0.83 AL/min/pmol CYP1A2. These findings suggest that CYP3A4 and CYP1A2 enzymes are major enzymes contributing to the metabolism of KR-32570.

  7. Single-enzyme kinetics with branched pathways: exact theory and series expansion

    CERN Document Server

    Garai, Ashok

    2014-01-01

    The progress of the successive rounds of catalytic conversion of substrates into product(s) by a single enzyme is characterized by the distribution of turnover times. Establishing the most general form of dependence of this distribution on the substrate concentration [S] is one of the fundamental challenges in single molecule enzymology. The distribution of the times of dwell of a molecular motor at the successive positions on its track is an analogous quantity. We derive approximate series expansions for the [ATP]-dependence of the first two moments of the dwell time distributions of motors that catalyze hydrolysis of ATP to draw input energy. Comparison between our results for motors with branched pathways and the corresponding expressions reported earlier for linear enzymatic pathways provides deep insight into the effects of the branches. Such insight is likely to help in discovering the most general form of [S]-dependence of these fundamental distributions.

  8. Polyol specificity of recombinant Arabidopsis thaliana sorbitol dehydrogenase studied by enzyme kinetics and in silico modeling

    Directory of Open Access Journals (Sweden)

    María Francisca eAguayo

    2015-02-01

    Full Text Available Polyols are enzymatically-produced plant compounds which can act as compatible solutes during periods of abiotic stress. NAD+-dependent SORBITOL DEHYDROGENASE (SDH, E.C. 1.1.1.14 from Arabidopsis thaliana L. (AtSDH is capable of oxidizing several polyols including sorbitol, ribitol and xylitol. In the present study, enzymatic assays using recombinant AtSDH demonstrated a higher specificity constant for xylitol compared to sorbitol and ribitol, all of which are C2 (S and C4 (R polyols. Enzyme activity was reduced by preincubation with ethylenediaminetetraacetic acid (EDTA, indicating a requirement for zinc ions. In humans, it has been proposed that sorbitol becomes part of a pentahedric coordination sphere of the catalytic zinc during the reaction mechanism. In order to determine the validity of this pentahedric coordination model in a plant SDH, homology modeling and Molecular Dynamics simulations of AtSDH ternary complexes with the three polyols were performed using crystal structures of human and Bemisia argentifolii (Genn. (Hemiptera: Aleyrodidae SDHs as scaffolds. The results indicate that the differences in interaction with structural water molecules correlate very well with the observed enzymatic parameters, validate the proposed pentahedric coordination of the catalytic zinc ion in a plant SDH, and provide an explanation for why AtSDH shows a preference for polyols with a chirality of C2 (S and C4 (R.

  9. Real-Time Enzyme Kinetics by Quantitative NMR Spectroscopy and Determination of the Michaelis-Menten Constant Using the Lambert-W Function

    Science.gov (United States)

    Her, Cheenou; Alonzo, Aaron P.; Vang, Justin Y.; Torres, Ernesto; Krishnan, V. V.

    2015-01-01

    Enzyme kinetics is an essential part of a chemistry curriculum, especially for students interested in biomedical research or in health care fields. Though the concept is routinely performed in undergraduate chemistry/biochemistry classrooms using other spectroscopic methods, we provide an optimized approach that uses a real-time monitoring of the…

  10. Developments in Analytical Chemistry: Acoustically Levitated Drop Reactors for Enzyme Reaction Kinetics and Single-Walled Carbon Nanotube-Based Sensors for Detection of Toxic Organic Phosphonates

    Science.gov (United States)

    Field, Christopher Ryan

    2009-01-01

    Developments in analytical chemistry were made using acoustically levitated small volumes of liquid to study enzyme reaction kinetics and by detecting volatile organic compounds in the gas phase using single-walled carbon nanotubes. Experience gained in engineering, electronics, automation, and software development from the design and…

  11. Kinetic and mutational dissection of the two ATPase activities of terminase, the DNA packaging enzyme of bacteriophage Chi.

    Science.gov (United States)

    Hwang, Y; Catalano, C E; Feiss, M

    1996-02-27

    Terminase the DNA packaging enzyme of bacteriophage chi, is a heteromultimer of gpNul (21 kDa) and gpA (74 kDa) subunits, encoded by the chi Nul and A genes, respectively. Sequence comparisons indicate that both gpNu1 and gpA have a match to the P-loop motif of ATPase centers, which is a glycine-rich segment followed by a lysine. By site-specific mutagenesis, we changed the lysines of the putative P-loops of gpNul (k35) and gpA (K497) to arginine, alanine, or aspartic acid, and studied the mutant enzymes by kinetic analysis and photochemical cross-linking with 8-azido-ATP. Both the gpNul and gpA subunits of wild-type terminase were covalently modified with 8-N3[32P] ATP in the presence of UV light. Saturation occurred with apparent dissociation constants of 508 and 3.5 microM for gpNul and gpA, resepctively. ATPase assays showed two activities: a low-affinity activity (Km=469 microM), and a high-affinity activity (Km=4.6 microM). The gpNul K35A and gpNul K35D mutant terminases showed decreased activity in the low-affinity ATPase activity. The reduced activities of these enzymes were recovered when 10 times more DNA was added, suggesting that the primary defect of the enzymes is alteration of the nonspecific, double-stranded DNA binding activity of terminase. ATPase assays and photolabeling of the gpA K497A and gpA K497D mutant terminases showed reduced affinity for ATP at the high-affinity site which was not restored by increased DNA. In summary, the results indicate the presence of a low-affinity, DNA-stimulated ATPase center in gpNul, and a high-affinity site in gpA.

  12. The effect of organic solvents on enzyme kinetic parameters of human CYP3A4 and CYP1A2 in vitro.

    Science.gov (United States)

    Rokitta, Dennis; Pfeiffer, Kay; Streich, Christina; Gerwin, Henrik; Fuhr, Uwe

    2013-10-01

    Abstract Enzyme kinetic parameters provide essential quantitative information about characterization of individual steps in drug metabolism. Such enzymes are located in a (partially) aqueous environment. For in vitro measurements potential lipophilic substrates regularly require organic solvents to achieve concentrations sufficient for access of the drug to the binding site of the enzyme. However, solvents may interact with the enzymes. In this study, we investigated the effects of methanol, ethanol, acetonitrile and dimethyl sulfoxide (1% to 4%) on the assessment of km, Vmax and Clint for the metabolism of midazolam via CYP3A4 to 1-hydroxymidazolam and the metabolism of caffeine to paraxanthine via CYP1A2 using expressed enzymes in vitro. The presence of acetonitrile proved the highest apparent Vmax value for paraxanthine formation but the lowest values for 1-hydroxymidazolam formation. The km value for midazolam showed no systematic effects of organic solvents, while for caffeine km was up to 8-fold lower for solvent free samples compared to solvent containing samples. The present example suggests that effects of solvents may considerably influence enzyme kinetic parameters beyond a mere change in apparent activity. These effects illustrate a difference for individual enzyme--substrate pairs, solvents, and solvent concentrations. What remains is the determination to which extent these effects compromise in vitro-in vivo extrapolations, and which solvents are most appropriate.

  13. Kinetic modelling of coupled transport across biological membranes.

    Science.gov (United States)

    Korla, Kalyani; Mitra, Chanchal K

    2014-04-01

    In this report, we have modelled a secondary active co-transporter (symport and antiport), based on the classical kinetics model. Michaelis-Menten model of enzyme kinetics for a single substrate, single intermediate enzyme catalyzed reaction was proposed more than a hundred years ago. However, no single model for the kinetics of co-transport of molecules across a membrane is available in the literature We have made several simplifying assumptions and have followed the basic Michaelis-Menten approach. The results have been simulated using GNU Octave. The results will be useful in general kinetic simulations and modelling.

  14. Off-line form of the Michaelis-Menten equation for studying the reaction kinetics in a polymer microchip integrated with enzyme microreactor.

    Science.gov (United States)

    Liu, Ai-Lin; Zhou, Ting; He, Feng-Yun; Xu, Jing-Juan; Lu, Yu; Chen, Hong-Yuan; Xia, Xing-Hua

    2006-06-01

    We firstly transformed the traditional Michaelis-Menten equation into an off-line form which can be used for evaluating the Michaelis-Menten constant after the enzymatic reaction. For experimental estimation of the kinetics of enzymatic reactions, we have developed a facile and effective method by integrating an enzyme microreactor into direct-printing polymer microchips. Strong nonspecific adsorption of proteins was utilized to effectively immobilize enzymes onto the microchannel wall, forming the integrated on-column enzyme microreactor in a microchip. The properties of the integrated enzyme microreactor were evaluated by using the enzymatic reaction of glucose oxidase (GOx) with its substrate glucose as a model system. The reaction product, hydrogen peroxide, was electrochemically (EC) analyzed using a Pt microelectrode. The data for enzyme kinetics using our off-line form of the Michaelis-Menten equation was obtained (K(m) = 2.64 mM), which is much smaller than that reported in solution (K(m) = 6.0 mM). Due to the hydrophobic property and the native mesoscopic structure of the poly(ethylene terephthalate) film, the immobilized enzyme in the microreactor shows good stability and bioactivity under the flowing conditions.

  15. The Non-Linear Child: Ontogeny, Isoniazid Concentration, and NAT2 Genotype Modulate Enzyme Reaction Kinetics and Metabolism

    Directory of Open Access Journals (Sweden)

    Zoe Rogers

    2016-09-01

    Full Text Available N-acetyltransferase 2 (NAT2 catalyzes the acetylation of isoniazid to N-acetylisoniazid. NAT2 polymorphism explains 88% of isoniazid clearance variability in adults. We examined the effects of clinical and genetic factors on Michaelis-Menten reaction kinetic constants of maximum velocity (Vmax and affinity (Km in children 0–10 years old. We measured the rates of isoniazid elimination and N-acetylisoniazid production in the blood of 30 children. Since maturation effects could be non-linear, we utilized a pharmacometric approach and the artificial intelligence method, multivariate adaptive regression splines (MARS, to identify factors predicting NAT2 Vmax and Km by examining clinical, genetic, and laboratory factors in toto. Isoniazid concentration predicted both Vmax and Km and superseded the contribution of NAT2 genotype. Age non-linearly modified the NAT2 genotype contribution until maturation at ≥5.3 years. Thus, enzyme efficiency was constrained by substrate concentration, genes, and age. Since MARS output is in the form of basis functions and equations, it allows multiscale systems modeling from the level of cellular chemical reactions to whole body physiological parameters, by automatic selection of significant predictors by the algorithm.

  16. Selective production of 1-monocaprin by porcine liver carboxylesterase-catalyzed esterification: Its enzyme kinetics and catalytic performance.

    Science.gov (United States)

    Park, Kyung-Min; Lee, Jong-Hyuk; Hong, Sung-Chul; Kwon, Chang Woo; Jo, Minje; Choi, Seung Jun; Kim, Keesung; Chang, Pahn-Shick

    2016-01-01

    Porcine liver carboxylesterase (PLE) belongs to carboxylesterase family (EC 3.1.1.1) as a serine-type esterase. The PLE-catalyzed esterification of capric acid with glycerol in reverse micelles was investigated on the catalytic performance and enzyme kinetics. The most suitable structure of reverse micelles was comprised of isooctane (reaction medium) and bis(2-ethylhexyl) sodium sulfosuccinate (AOT, anionic surfactant) with 0.1 of R-value ([water]/[surfactant]) and 3.0 of G/F-value ([glycerol]/[fatty acid]) for the PLE-catalyzed esterification. In the aspect of regio-selectivity, the PLE mainly produced 1-monocaprin without any other products (di- and/or tricaprins of subsequent reactions). Furthermore, the degree of esterification at equilibrium state (after 4 h from the initiation) was 62.7% under the optimum conditions at pH 7.0 and 60 °C. Based on Hanes-Woolf plot, the apparent Km and Vmax values were calculated to be 16.44 mM and 38.91 μM/min/mg protein, respectively.

  17. Application of robotics to steady state enzyme kinetics: analysis of tight-binding inhibitors of dipeptidyl peptidase IV.

    Science.gov (United States)

    Wang, Aiying; Huang, Yanting; Taunk, Prakash; Magnin, David R; Ghosh, Krishnendu; Robertson, James G

    2003-10-15

    Using available commercial robotics and instrumentation, we developed a fully automated and rigorous steady state enzyme kinetic assay for dipeptidyl peptidase IV (DPP IV; E.C. 3.4.14.5). The automated assay was validated with isoleucyl thiazolidide, a potent inhibitor of DPP IV with K(is)=110nM. Signal window analysis indicated that the assay had a 98% probability of detecting an inhibitor yielding 15% inhibition, with a predicted false positive rate of 0.13%. A mechanistic inhibition version of the automated assay was validated with isoleucyl 4-cyanothiazolidide, a very potent inhibitor of DPP IV. Isoleucyl 4-cyanothiazolidide was a competitive inhibitor of purified porcine DPP IV with K(is)=1 nM. Similar K(is) values were obtained for purified rat DPP IV and for DPP IV activity in human plasma from normal and diabetic donors. The pH dependence of K(is) for isoleucyl 4-cyanothiazolidide yielded a bell-shaped profile, with pK(a)=5.0 and pK(b)=7.6. To date, over 100,000 data points have been generated in profiling targeted compound libraries and in the analysis of tight-binding inhibitors of DPP IV. The data also show that robotic analysis is capable of producing full mechanistic inhibition analysis in a timely fashion to support drug discovery.

  18. Discrimination among eight modified michaelis-menten kinetics models of cellulose hydrolysis with a large range of substrate/enzyme ratios: inhibition by cellobiose.

    Science.gov (United States)

    Bezerra, Rui M F; Dias, Albino A

    2004-03-01

    The kinetics of exoglucanase (Cel7A) from Trichoderma reesei was investigated in the presence of cellobiose and 24 different enzyme/Avicel ratios for 47 h, in order to establish which of the eight available kinetic models best explained the factors involved. The heterogeneous catalysis was studied and the kinetic parameters were estimated employing integrated forms of Michaelis-Menten equations through the use of nonlinear least squares. It was found that cellulose hydrolysis follows a model that takes into account competitive inhibition by cellobiose (final product) with the following parameters: Km = 3.8 mM, Kic = 0.041 mM, kcat = 2 h-1 (5.6 x 10-4 s-1). Other models, such as mixed type inhibition and those incorporating improvements concerning inhibition by substrate and parabolic inhibition, increased the modulation performance very slightly. The results support the hypothesis that nonproductive enzyme substrate complexes, parabolic inhibition, and enzyme inactivation (Selwyn test) are not the principal constraints in enzymatic cellulose hydrolysis. Under our conditions, the increment in hydrolysis was not significant for substrate/enzyme ratios <6.5.

  19. Numerical stability of finite difference algorithms for electrochemical kinetic simulations. Matrix stability analysis of the classic explicit, fully implicit and Crank-Nicolson methods, extended to the 3- and 4-point gradient approximation at the electrodes

    DEFF Research Database (Denmark)

    Bieniasz, Leslaw K.; Østerby, Ole; Britz, Dieter

    1995-01-01

    We extend the analysis of the stepwise numerical stability of the classic explicit, fully implicit and Crank-Nicolson finite difference algorithms for electrochemical kinetic simulations, to the multipoint gradient approximations at the electrode. The discussion is based on the matrix method...... of stability analysis....

  20. Spectroscopic and Kinetic Characterization of Peroxidase-Like π-Cation Radical Pinch-Porphyrin-Iron(III Reaction Intermediate Models of Peroxidase Enzymes

    Directory of Open Access Journals (Sweden)

    Samuel Hernández Anzaldo

    2016-06-01

    Full Text Available The spectroscopic and kinetic characterization of two intermediates from the H2O2 oxidation of three dimethyl ester [(proto, (meso, (deuteroporphyrinato (picdien]Fe(III complexes ([FePPPic], [FeMPPic] and [FeDPPic], respectively pinch-porphyrin peroxidase enzyme models, with s = 5/2 and 3/2 Fe(III quantum mixed spin (qms ground states is described herein. The kinetic study by UV/Vis at λmax = 465 nm showed two different types of kinetics during the oxidation process in the guaiacol test for peroxidases (1–3 + guaiacol + H2O2 → oxidation guaiacol products. The first intermediate was observed during the first 24 s of the reaction. When the reaction conditions were changed to higher concentration of pinch-porphyrins and hydrogen peroxide only one type of kinetics was observed. Next, the reaction was performed only between pinch-porphyrins-Fe(III and H2O2, resulting in only two types of kinetics that were developed during the first 0–4 s. After this time a self-oxidation process was observed. Our hypotheses state that the formation of the π-cation radicals, reaction intermediates of the pinch-porphyrin-Fe(III family with the ligand picdien [N,N’-bis-pyridin-2-ylmethyl-propane-1,3-diamine], occurred with unique kinetics that are different from the overall process and was involved in the oxidation pathway. UV-Vis, 1H-NMR and ESR spectra confirmed the formation of such intermediates. The results in this paper highlight the link between different spectroscopic techniques that positively depict the kinetic traits of artificial compounds with enzyme-like activity.

  1. The comparison of the estimation of enzyme kinetic parameters by fitting reaction curve to the integrated Michaelis-Menten rate equations of different predictor variables.

    Science.gov (United States)

    Liao, Fei; Zhu, Xiao-Yun; Wang, Yong-Mei; Zuo, Yu-Ping

    2005-01-31

    The estimation of enzyme kinetic parameters by nonlinear fitting reaction curve to the integrated Michaelis-Menten rate equation ln(S(0)/S)+(S(0)-S)/K(m)=(V(m)/K(m))xt was investigated and compared to that by fitting to (S(0)-S)/t=V(m)-K(m)x[ln(S(0)/S)/t] (Atkins GL, Nimmo IA. The reliability of Michaelis-Menten constants and maximum velocities estimated by using the integrated Michaelis-Menten equation. Biochem J 1973;135:779-84) with uricase as the model. Uricase reaction curve was simulated with random absorbance error of 0.001 at 0.075 mmol/l uric acid. Experimental reaction curve was monitored by absorbance at 293 nm. For both CV and deviation kinetic parameters and applicable for the characterization of enzyme inhibitors.

  2. Fundamental study of the mechanism and kinetics of cellulose hydrolysis by acids and enzymes. Progress report, June 1, 1975--December 31, 1975

    Energy Technology Data Exchange (ETDEWEB)

    Tsao, G T

    1976-02-01

    This project deals with acidic and enzymatic hydrolysis of cellulosic materials. Highlights of the first eight months of out project are as follows. (1) Essentially homogeneous C/sub 1/, C/sub x/, and Cellobiase enzyme have been isolated from Cellulase-Onozuka of Trichodeima viride origin. Besides the 3 major components, one protein of a molecular weight of 10,000 was purified, which has strong C/sub x/ activity and also another (M.W. 23,000) with strong C/sub 1/ activity. (2) Kinetics of Cellobiase has been investigated and its kinetics constant accurately determined. Immobilization of this enzyme on porous glass was successful. (3) Absorption of SO/sub 2/ at atmospheric pressure increased digestibility of delignated cellulose but not the natural materials such as corn stalk. A pressurizable absorption unit is being built. (4) Kinetics models for purified C/sub 1/ and C/sub x/ are postulated and equations derived. Experimental tests will be made when sufficient quantities of purified C/sub 1/ and C/sub x/ enzyme are prepared. (5) A lignin digesting, white-rot fungus, Pleurotus ostreatus, has cultivated in our laboratory. It has grown well and heavy. (6) A number of electron micrographs were made with untreated cellulosic materials, which showed that the method of drying the specimens for electromicroscopy is important. Ordinary drying procedures will alter their physical structures. A technique, critical point drying, is being practiced, which will not change the structure. (auth)

  3. Occurrence of dead core in catalytic particles containing immobilized enzymes: analysis for the Michaelis-Menten kinetics and assessment of numerical methods.

    Science.gov (United States)

    Pereira, Félix Monteiro; Oliveira, Samuel Conceição

    2016-11-01

    In this article, the occurrence of dead core in catalytic particles containing immobilized enzymes is analyzed for the Michaelis-Menten kinetics. An assessment of numerical methods is performed to solve the boundary value problem generated by the mathematical modeling of diffusion and reaction processes under steady state and isothermal conditions. Two classes of numerical methods were employed: shooting and collocation. The shooting method used the ode function from Scilab software. The collocation methods included: that implemented by the bvode function of Scilab, the orthogonal collocation, and the orthogonal collocation on finite elements. The methods were validated for simplified forms of the Michaelis-Menten equation (zero-order and first-order kinetics), for which analytical solutions are available. Among the methods covered in this article, the orthogonal collocation on finite elements proved to be the most robust and efficient method to solve the boundary value problem concerning Michaelis-Menten kinetics. For this enzyme kinetics, it was found that the dead core can occur when verified certain conditions of diffusion-reaction within the catalytic particle. The application of the concepts and methods presented in this study will allow for a more generalized analysis and more accurate designs of heterogeneous enzymatic reactors.

  4. Interaction between wheat alpha-amylase/trypsin bi-functional inhibitor and mammalian digestive enzymes: Kinetic, equilibrium and structural characterization of binding.

    Science.gov (United States)

    Cuccioloni, Massimiliano; Mozzicafreddo, Matteo; Ali, Ishtiaq; Bonfili, Laura; Cecarini, Valentina; Eleuteri, Anna Maria; Angeletti, Mauro

    2016-12-15

    Alpha-amylase/trypsin bi-functional inhibitors (ATIs) are non-gluten protein components of wheat and other cereals that can hypersensitise the human gastrointestinal tract, eventually causing enteropathies in predisposed individuals. These inhibitory proteins can act both directly by targeting specific pro-inflammatory receptors, and indirectly by impairing the activity of digestive enzymes, the latter event causing the accumulation of undigested peptides with potential immunogenic properties. Herein, according to a concerted approach based on in vitro and in silico methods we characterized kinetics, equilibrium parameters and modes of binding of the complexes formed between wheat ATI and two representative mammalian digestive enzymes, namely trypsin and alpha-amylase. Interestingly, we demonstrated ATI to target both enzymes with independent binding sites and with moderately high affinity.

  5. Effect of pulsed electric field treatment on enzyme kinetics and thermostability of endogenous ascorbic acid oxidase in carrots (Daucus carota cv. Nantes).

    Science.gov (United States)

    Leong, Sze Ying; Oey, Indrawati

    2014-03-01

    The objective of this research was to study the enzyme kinetics and thermostability of endogenous ascorbic acid oxidase (AAO) in carrot purée (Daucus carota cv. Nantes) after being treated with pulsed electric field (PEF) processing. Various PEF treatments using electric field strength between 0.2 and 1.2kV/cm and pulsed electrical energy between 1 and 520kJ/kg were conducted. The enzyme kinetics and the kinetics of AAO thermal inactivation (55-70°C) were described using Michaelis-Menten model and first order reaction model, respectively. Overall, the estimated Vmax and KM values were situated in the same order of magnitude as the untreated carrot purée after being exposed to pulsed electrical energy between 1 and 400kJ/kg, but slightly changed at pulsed electrical energy above 500kJ/kg. However, AAO presented different thermostability depending on the electric field strength applied. After PEF treatment at the electric field strength between 0.2 and 0.5kV/cm, AAO became thermolabile (i.e. increase in inactivation rate (k value) at reference temperature) but the temperature dependence of k value (Ea value) for AAO inactivation in carrot purée decreased, indicating that the changes in k values were less temperature dependent. It is obvious that PEF treatment affects the temperature stability of endogenous AAO. The changes in enzyme kinetics and thermostability of AAO in carrot purée could be related to the resulting carrot purée composition, alteration in intracellular environment and the effective concentration of AAO released after being subjected to PEF treatment.

  6. Kinetic characterization of glucose aerodehydrogenase from Aspergillus niger EMS-150-F after optimizing the dose of mutagen for enhanced production of enzyme.

    Science.gov (United States)

    Umbreen, Huma; Zia, Muhammad Anjum; Rasul, Samreen

    2013-12-01

    In the present study enhanced production of glucose aerodehydrogenase from Aspergillus niger has been achieved after optimizing the dose of chemical mutagen ethyl methane sulfonate (EMS) that has not been reported earlier. Different doses of mutagen were applied and a strain was developed basing upon the best production. The selected strain Aspergillus niger EMS-150-F was optimized for nutrient requirements in order to produce enzyme through fermentation and the results showed the best yield at 2% corn steep liquor (CSL), 36 hours fermentation time, pH 5, 30 °C temperature, 0.3% KH2PO4, 0.3% urea and 0.06% CaCO3. The enzyme was then purified and resulted in 57.88 fold purification with 52.12% recovery. On kinetic characterization, the enzyme showed optimum activity at pH 6 and temperature 30 °C. The Michaelis-Menton constants (K(m), Vmax, Kcat and Kcat/K(m)) were 20 mM, 45.87 U mL(-1), 1118.81 s(-1) and 55.94 s(-1) mM(-1), respectively. The enzyme was found to be thermally stable and the enthalpy and free energy showed an increase with increase in temperature and ΔS* was highly negative proving the enzyme from A. niger EMS-150-F resistant to temperature and showing a very little disorderliness.

  7. Implication for functions of the ectopic adipocyte copper amine oxidase (AOC3 from purified enzyme and cell-based kinetic studies.

    Directory of Open Access Journals (Sweden)

    Sam H Shen

    Full Text Available AOC3 is highly expressed in adipocytes and smooth muscle cells, but its function in these cells is currently unknown. The in vivo substrate(s of AOC3 is/are also unknown, but could provide an invaluable clue to the enzyme's function. Expression of untagged, soluble human AOC3 in insect cells provides a relatively simple means of obtaining pure enzyme. Characterization of enzyme indicates a 6% titer for the active site 2,4,5-trihydroxyphenylalanine quinone (TPQ cofactor and corrected k(cat values as high as 7 s(-1. Substrate kinetic profiling shows that the enzyme accepts a variety of primary amines with different chemical features, including nonphysiological branched-chain and aliphatic amines, with measured k(cat/K(m values between 10(2 and 10(4 M(-1 s(-1. K(m(O(2 approximates the partial pressure of oxygen found in the interstitial space. Comparison of the properties of purified murine to human enzyme indicates k(cat/K(m values that are within 3 to 4-fold, with the exception of methylamine and aminoacetone that are ca. 10-fold more active with human AOC3. With drug development efforts investigating AOC3 as an anti-inflammatory target, these studies suggest that caution is called for when screening the efficacy of inhibitors designed against human enzymes in non-transgenic mouse models. Differentiated murine 3T3-L1 adipocytes show a uniform distribution of AOC3 on the cell surface and whole cell K(m values that are reasonably close to values measured using purified enzymes. The latter studies support a relevance of the kinetic parameters measured with isolated AOC3 variants to adipocyte function. From our studies, a number of possible substrates with relatively high k(cat/K(m have been discovered, including dopamine and cysteamine, which may implicate a role for adipocyte AOC3 in insulin-signaling and fatty acid metabolism, respectively. Finally, the demonstrated AOC3 turnover of primary amines that are non-native to human tissue suggests

  8. Substrate specificity and kinetic properties of enzymes belonging to the hormone-sensitive lipase family: comparison with non-lipolytic and lipolytic carboxylesterases.

    Science.gov (United States)

    Chahinian, Henri; Ali, Yassine Ben; Abousalham, Abdelkarim; Petry, Stefan; Mandrich, Luigi; Manco, Guiseppe; Canaan, Stephane; Sarda, Louis

    2005-12-30

    We have studied the kinetics of hydrolysis of triacylglycerols, vinyl esters and p-nitrophenyl butyrate by four carboxylesterases of the HSL family, namely recombinant human hormone-sensitive lipase (HSL), EST2 from Alicyclobacillus acidocaldarius, AFEST from Archeoglobus fulgidus, and protein RV1399C from Mycobacterium tuberculosis. The kinetic properties of enzymes of the HSL family have been compared to those of a series of lipolytic and non-lipolytic carboxylesterases including human pancreatic lipase, guinea pig pancreatic lipase related protein 2, lipases from Mucor miehei and Thermomyces lanuginosus, cutinase from Fusarium solani, LipA from Bacillus subtilis, porcine liver esterase and Esterase A from Aspergilus niger. Results indicate that human HSL, together with other lipolytic carboxylesterases, are active on short chain esters and hydrolyze water insoluble trioctanoin, vinyl laurate and olive oil, whereas the action of EST2, AFEST, protein RV1399C and non-lipolytic carboxylesterases is restricted to solutions of short chain substrates. Lipolytic and non-lipolytic carboxylesterases can be differentiated by their respective value of K(0.5) (apparent K(m)) for the hydrolysis of short chain esters. Among lipolytic enzymes, those possessing a lid domain display higher activity on tributyrin, trioctanoin and olive oil suggesting, then, that the lid structure contributes to enzyme binding to triacylglycerols. Progress reaction curves of the hydrolysis of p-nitrophenyl butyrate by lipolytic carboxylesterases with lid domain show a latency phase which is not observed with human HSL, non-lipolytic carboxylesterases, and lipolytic enzymes devoid of a lid structure as cutinase.

  9. Multistep generalized transformation method applied to solving equations of discrete and continuous time-fractional enzyme kinetics

    Science.gov (United States)

    Vosika, Z.; Mitić, V. V.; Vasić, A.; Lazović, G.; Matija, L.; Kocić, Lj. M.

    2017-03-01

    In this paper, Caputo based Michaelis-Menten kinetic model based on Time Scale Calculus (TSC) is proposed. The main reason for its consideration is a study of tumor cells population growth dynamics. In the particular case discrete-continuous time kinetics, Michaelis-Menten model is numerically treated, using a new algorithm proposed by authors, called multistep generalized difference transformation method (MSGDETM). In addition numerical simulations are performed and is shown that it represents the upgrade of the multi-step variant of generalized differential transformation method (MSGDTM). A possible conditions for its further development are discussed and possible experimental verification is described.

  10. Study on the kinetics of saccharifying enzymes of Luzhou-flavor daqu%浓香型大曲糖化动力学研究

    Institute of Scientific and Technical Information of China (English)

    任飞; 张晓宇

    2013-01-01

    In order to study on the kinetics of saccharifying enzymes of Luzhou-flavor Daqu, using the enzymes of 5% Daqu hydrolyzed soluble starch, at pH 4. 6, 30℃ , an Michaelis-Menten equation was deduced. The optimum pH value for the catalysis of saccharifying enzymes of Luzhou-flavor Daqu was 6. 0. Studying the catalytic reaction rate under different temperatures, the calculation results showed that the activation energy of the enzyme catalytic reaction was 40. 966 kJ/mol. The kinetics of saccharifying enzymes of Luzhou-fla-vor Daqu was similar to a single enzyme, During the production of Luzhou-flavor liquor, it will contribute to manage the saccharification and fermentation process simultaneously by controlling the conditions of pits input into the cellar, such as the concentration of starch and starter, initial acidity and temperature.%为研究浓香型大曲的糖化动力学,用5%的大曲浸出液水解可溶性淀粉,在pH4.6、30℃的条件下,用菲林试剂法测定还原糖的含量,得出米氏方程;浓香型大曲糖化反应的最适pH值为6.0;在不同温度下测定糖化反应速率,计算得到活化能E为40.966 kJ/mol.试验证明,浓香型大曲糖化酶类的糖化反应动力学与单一酶相似.通过控制淀粉浓度(投粮量)、用曲量、入窖酸度和温度可以使浓香型大曲酒的糖化、发酵过程协调进行.

  11. Kinetic properties of two Rhizopus exo-polygalacturonase enzymes hydrolyzing galacturonic acid oligomers using isothermal titration calorimetry

    Science.gov (United States)

    The kinetic characteristics of two Rhizopus oryzae exo-polygalacturonases acting on galacturonic acid oligomers (GalpA) were determined using isothermal titration calorimetry (ITC). RPG15 hydrolyzing (GalpA)2 demonstrated a Km of 55 uM and kcat of 10.3 s^-1^ while RPG16 was shown to have greater af...

  12. The reaction kinetics of 3-hydroxybenzoate 6-hydroxylase from Rhodococcus jostii RHA1 provide an understanding of the para-hydroxylation enzyme catalytic cycle.

    Science.gov (United States)

    Sucharitakul, Jeerus; Tongsook, Chanakan; Pakotiprapha, Danaya; van Berkel, Willem J H; Chaiyen, Pimchai

    2013-12-06

    3-Hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1 is an NADH-specific flavoprotein monooxygenase that catalyzes the para-hydroxylation of 3-hydroxybenzoate (3HB) to form 2,5-dihydroxybenzoate (2,5-DHB). Based on results from stopped-flow spectrophotometry, the reduced enzyme-3HB complex reacts with oxygen to form a C4a-peroxy flavin with a rate constant of 1.13 ± 0.01 × 10(6) m(-1) s(-1) (pH 8.0, 4 °C). This intermediate is subsequently protonated to form a C4a-hydroperoxyflavin with a rate constant of 96 ± 3 s(-1). This step shows a solvent kinetic isotope effect of 1.7. Based on rapid-quench measurements, the hydroxylation occurs with a rate constant of 36 ± 2 s(-1). 3HB6H does not exhibit substrate inhibition on the flavin oxidation step, a common characteristic found in most ortho-hydroxylation enzymes. The apparent kcat at saturating concentrations of 3HB, NADH, and oxygen is 6.49 ± 0.02 s(-1). Pre-steady state and steady-state kinetic data were used to construct the catalytic cycle of the reaction. The data indicate that the steps of product release (11.7 s(-1)) and hydroxylation (36 ± 2 s(-1)) partially control the overall turnover.

  13. Kinetics and molecular docking studies of the inhibitions of angiotensin converting enzyme and renin activities by hemp seed (Cannabis sativa L.) peptides.

    Science.gov (United States)

    Girgih, Abraham T; He, Rong; Aluko, Rotimi E

    2014-05-07

    Four novel peptide sequences (WVYY, WYT, SVYT, and IPAGV) identified from an enzymatic digest of hemp seed proteins were used for enzyme inhibition kinetics and molecular docking studies. Results showed that WVYY (IC50 = 0.027 mM) was a more potent (p < 0.05) ACE-inhibitory peptide than WYT (IC50 = 0.574 mM). However, WYT (IC50 = 0.054 mM) and SVYT (IC50 = 0.063 mM) had similar renin-inhibitory activity, which was significantly better than that of IPAGV (IC50 = 0.093 mM). Kinetics studies showed that WVYY had a lower inhibition constant (Ki) of 0.06 mM and hence greater affinity for ACE when compared to the 1.83 mM obtained for WYT. SVYT had lowest Ki value of 0.89 mM against renin, when compared to the values obtained for WYT and IPAGV. Molecular docking results showed that the higher inhibitory activities of WVYY and SVYT were due to the greater degree of noncovalent bond-based interactions with the enzyme protein, especially formation of higher numbers of hydrogen bonds with active site residues.

  14. Production of manganese peroxidase and laccase in a solid-state bioreactor and modeling of enzyme production kinetics.

    Science.gov (United States)

    Moilanen, Ulla; Winquist, Erika; Mattila, Tuomas; Hatakka, Annele; Eerikäinen, Tero

    2015-01-01

    Lignin-modifying enzymes have various promising applications such as biobleaching, biopulping, the functionalization of lignocellulosic materials, the modification of wood fibers, the remediation of contaminated soil and effluents, as well as improvement of the enzymatic hydrolysis of lignocellulosic substrates. In this study, the production of laccase and manganese peroxidase (MnP) in solid-state cultivation was examined. Oat husks were used as an inexpensive substrate for the white-rot fungus Cerrena unicolor PM170798 (FBCC 387). The addition of a fines fraction (consisting of oat flour and finely ground husks) enhanced MnP production fivefold and laccase production almost threefold. The enzyme production was studied first on a 100 g scale, and the cultivation experiments were then repeated at a larger laboratory-scale (4 kg) in a solid-state bioreactor. High enzyme activity levels were obtained (MnP: 340 nkat g(-1) DM, laccase: 470 nkat g(-1) DM). In addition, the correlation between the CO2 evolution rate and enzyme production was mathematically modeled from the bioreactor experimental data. The model parameters could be used to predict enzyme production.

  15. Influence of temperature and solvent concentration on the kinetics of the enzyme carbonic anhydrase in carbon capture technology

    DEFF Research Database (Denmark)

    Gladis, Arne; Deslauriers, Maria Gundersen; Fosbøl, Philip Loldrup

    2017-01-01

    In this study the effect of carbonic anhydrase addition on the absorption of CO2 was investigated in a wetted wall column apparatus. Four different solvents: the primary amine monoethanolamine (MEA), the sterically hindered primary amine 2-amino-2-methyl-1-propanol (AMP), the tertiary amine N......-methyl-diethanolamine (MDEA) and the carbonate salt solution K2CO3 were compared in concentrations from 5 to 50 wt% in a temperature range of 298–328 K with and without enzyme. Necessary mass transfer parameters such as liquid side mass transfer coefficient and solvent and enzyme reaction rates were determined...... and benchmarked to a 30 wt% MEA solution. The study reveals that the addition of the enzyme carbonic anhydrase (CA) dramatically increases the liquid side mass transfer coefficient for MDEA, and K2CO3; AMP has a moderate increase whereas MEA was unchanged. The results confirm that just bicarbonate forming systems...

  16. Adsorption of Trametes versicolor laccase to soil iron and aluminum minerals: enzyme activity, kinetics and stability studies.

    Science.gov (United States)

    Wu, Yue; Jiang, Ying; Jiao, Jiaguo; Liu, Manqiang; Hu, Feng; Griffiths, Bryan S; Li, Huixin

    2014-02-01

    Laccases play an important role in the degradation of soil phenol or phenol-like substance and can be potentially used in soil remediation through immobilization. Iron and aluminum minerals can adsorb extracellular enzymes in soil environment. In the present study, we investigated the adsorptive interaction of laccase, from the white-rot fungus Trametes versicolor, with soil iron and aluminum minerals and characterized the properties of the enzyme after adsorption to minerals. Results showed that both soil iron and aluminum minerals adsorbed great amount of laccase, independent of the mineral specific surface areas. Adsorbed laccases retained 26-64% of the activity of the free enzyme. Compared to the free laccase, all adsorbed laccases showed higher Km values and lower Vmax values, indicating a reduced enzyme-substrate affinity and a lower rate of substrate conversion in reactions catalyzed by the adsorbed laccase. Adsorbed laccases exhibited increased catalytic activities compared to the free laccase at low pH, implying the suitable application of iron and aluminum mineral-adsorbed T. versicolor laccase in soil bioremediation, especially in acid soils. In terms of the thermal profiles, adsorbed laccases showed decreased thermal stability and higher temperature sensitivity relative to the free laccase. Moreover, adsorption improved the resistance of laccase to proteolysis and extended the lifespan of laccase. Our results implied that adsorbed T. versicolor laccase on soil iron and aluminum minerals had promising potential in soil remediation.

  17. Red Seaweed Enzyme-Catalyzed Bromination of Bromophenol Red: An Inquiry-Based Kinetics Laboratory Experiment for Undergraduates

    Science.gov (United States)

    Jittam, Piyachat; Boonsiri, Patcharee; Promptmas, Chamras; Sriwattanarothai, Namkang; Archavarungson, Nattinee; Ruenwongsa, Pintip; Panijpan, Bhinyo

    2009-01-01

    Haloperoxidase enzymes are of interest for basic and applied bioscientists because of their increasing importance in pharmaceutical industry and environmental cleanups. In a guided inquiry-based laboratory experiment for life-science, agricultural science, and health science undergraduates, the bromoperoxidase from a red seaweed was used to…

  18. Inhibitory and enzyme-kinetic investigation of chelerythrine and lupeol isolated from Zanthoxylum rhoifolium against krait snake venom acetylcholinesterase

    Energy Technology Data Exchange (ETDEWEB)

    Ahmad, Mustaq, E-mail: mushtaq213@yahoo.com [University of Science and Technology, Bannu, (Pakistan). Department of Biotechnology; Weber, Andrea D.; Zanon, Graciane; Tavares, Luciana de C.; Ilha, Vinicius; Dalcol, Ionara I.; Morel, Ademir F., E-mail: ademirfariasm@gmail.com [Universidade Federal de Santa Maria, RS (Brazil). Dept. de Quimica

    2014-01-15

    The in vitro activity of chelerythrine and lupeol, two metabolites isolated from Zanthoxylum rhoifolium were studied against the venom of the snake Bungarus sindanus (Elapidae). The venom, which is highly toxic to humans, consists mainly by the enzyme acetylcholinesterase (AChE). Both compounds showed activity against the venom, and the alkaloid chelerythrine presented higher activity than did triterpene lupeol. (author)

  19. Enzyme kinetics and distinct modulation of the protein kinase N family of kinases by lipid activators and small molecule inhibitors

    OpenAIRE

    Falk, Matthew D.; Liu, Wei; Bolaños, Ben; Unsal-Kacmaz, Keziban; Klippel, Anke; Grant, Stephan; Brooun, Alexei; Timofeevski, Sergei

    2014-01-01

    The PKN (protein kinase N) family of Ser/Thr protein kinases regulates a diverse set of cellular functions, such as cell migration and cytoskeletal organization. Inhibition of tumour PKN activity has been explored as an oncology therapeutic approach, with a PKN3-targeted RNAi (RNA interference)-derived therapeutic agent in Phase I clinical trials. To better understand this important family of kinases, we performed detailed enzymatic characterization, determining the kinetic mechanism and lipi...

  20. Enzyme transient state kinetics in crystal and solution from the perspective of a time-resolved crystallographer

    Directory of Open Access Journals (Sweden)

    Marius Schmidt

    2014-03-01

    Full Text Available With recent technological advances at synchrotrons [Graber et al., J. Synchrotron Radiat. 18, 658–670 (2011], it is feasible to rapidly collect time-resolved crystallographic data at multiple temperature settings [Schmidt et al., Acta Crystallogr. D 69, 2534–2542 (2013], from which barriers of activation can be extracted. With the advent of fourth generation X-ray sources, new opportunities emerge to investigate structure and dynamics of biological macromolecules in real time [M. Schmidt, Adv. Condens. Matter Phys. 2013, 1–10] in crystals and potentially from single molecules in random orientation in solution [Poon et al., Adv. Condens. Matter Phys. 2013, 750371]. Kinetic data from time-resolved experiments on short time-scales must be interpreted in terms of chemical kinetics [Steinfeld et al., Chemical Kinetics and Dynamics, 2nd ed. (Prentience Hall, 1985] and tied to existing time-resolved experiments on longer time-scales [Schmidt et al., Acta Crystallogr. D 69, 2534–2542 (2013; Jung et al., Nat. Chem. 5, 212–220 (2013]. With this article, we will review and outline steps that are required to routinely determine the energetics of reactions in biomolecules in crystal and solution with newest X-ray sources. In eight sections, we aim to describe concepts and experimental details that may help to inspire new approaches to collect and interpret these data.

  1. Functional expression of a fragment of human dihydroorotate dehydrogenase by means of the baculovirus expression vector system, and kinetic investigation of the purified recombinant enzyme.

    Science.gov (United States)

    Knecht, W; Bergjohann, U; Gonski, S; Kirschbaum, B; Löffler, M

    1996-08-15

    Human mitochondrial dihydroorotate dehydrogenase (the fourth enzyme of pyrimidine de novo synthesis) has been overproduced by means of a recombinant baculovirus that contained the human cDNA fragment for this protein. After virus infection and protein expression in Trichoplusia ni cells (BTI-Tn-5B1-4), the subcellular distribution of the recombinant dihydroorotate dehydrogenase was determined by two distinct enzyme-activity assays and by Western blot analysis with anti-(dihydroorotate dehydrogenase) Ig. The targeting of the recombinant protein to the mitochondria of the insect cells was verified. The activity of the recombinant enzyme in the mitochondria of infected cells was about 740-fold above the level of dihydroorotate dehydrogenase in human liver mitochondria. In a three-step procedure, dihydroorotate dehydrogenase was purified to a specific activity of greater than 50 U/mg. Size-exclusion chromatography showed a molecular mass of 42 kDa and confirmed the existence of the fully active enzyme as a monomeric species. Fluorimetric cofactor analysis revealed the presence of FMN in recombinant dihydroorotate dehydrogenase. By kinetics analysis, Km values for dihydroorotate and ubiquinone-50 were found to be 4 microM and 9.9 microM, respectively, while Km values for dihydroorotate and decylubiquinone were 9.4 microM and 13.7 microM, respectively. The applied expression system will allow preparation of large quantities of the enzyme for structure and function studies. Purified recombinant human dihytdroorotate dehydrogenase was tested for its sensitivity to a reported inhibitor A77 1726 (2-hydroxyethyliden-cyanoacetic acid 4-trifluoromethyl anilide), which is the active metabolite of the isoxazole derivative leflunomide [5-methyl-N-(4-trifluoromethyl-phenyl)-4-isoxazole carboximide]. An IC50 value of 1 microM was determined for A77 1726. Detailed kinetics experiments revealed uncompetitive inhibition with respect to dihydroorotate (Kiu = 0.94 microM) and non

  2. Novel mathematical models for cell-mediated cytotoxicity assays without applying enzyme kinetics but with combinations and probability: bystanders in bulk effector cells influence results of cell-mediated cytotoxicity assays.

    Science.gov (United States)

    Takayanagi, Toshiaki

    2011-07-01

    Cell-mediated cytotoxicity assays are widely implemented to evaluate cell-mediated cytotoxic activity, and some assays are analyzed using the analogy of enzyme kinetics. In the analogy, the effector cell is regarded as the enzyme, the target cell as the substrate, the effector cell-target cell conjugate as the enzyme-substrate complex and the dead target cell as the product. However, the assumptions analogous to those of enzyme kinetics are not always true in cell-mediated cytotoxicity assays, and the parameter analogous to the Michaelis-Menten constant is not constant but is dependent on the number of effector cells. Therefore I present novel mathematical models for cell-mediated cytotoxicity assays without applying enzyme kinetics. I instead use combinations and probability, because analysis of cell-mediated cytotoxicity assays by applying enzyme kinetics seems controversial. With my original models, I demonstrate simulations of the data in previously published papers. The results are exhibited in the same forms as the corresponding data. Comparing the simulation results with the published data, the results seem to agree well with the data. From simulations of cytotoxic assays with bulk effector cells, it appears that bystanders in bulk effector cells increase both the cytotoxic activity and the motility of effector cells.

  3. Classical antiparticles

    Energy Technology Data Exchange (ETDEWEB)

    Costella, J.P.; McKellar, B.H.J.; Rawlinson, A.A.

    1997-03-01

    We review how antiparticles may be introduced in classical relativistic mechanics, and emphasize that many of their paradoxical properties can be more transparently understood in the classical than in the quantum domain. (authors). 13 refs., 1 tab.

  4. Classical antiparticles

    CERN Document Server

    Costella, J P; Rawlinson, A A; Costella, John P.; Kellar, Bruce H. J. Mc; Rawlinson, Andrew A.

    1997-01-01

    We review how antiparticles may be introduced in classical relativistic mechanics, and emphasize that many of their paradoxical properties can be more transparently understood in the classical than in the quantum domain.

  5. Effect of morin on pharmacokinetics of piracetam in rats, in vitro enzyme kinetics and metabolic stability assay using rapid UPLC method.

    Science.gov (United States)

    Sahu, Kapendra; Shaharyar, Mohammad; Siddiqui, Anees A

    2013-07-01

    The aim of this study was to investigate the effect of Morin on the pharmacokinetics of Piracetam in rats, in vitro enzyme kinetics and metabolic stability (high throughput) studies using human liver microsomes in UPLC. For pharmacokinetics studies, male Wistar rats were pretreated with Morin (10 mg/kg) for one week and on the last day, a single dose of Piracetam (50 mg/kg) was given orally. In another group, both Morin and Piracetam were co-administered to evaluate the acute effect of Morin on Piracetam. The control group received oral distilled water for one week and administered with Piracetam on the last day. As Morin is an inhibitor of P- Glycoprotein (P-gp) and CYP 3A, it was anticipated to improve the bioavailability of Piracetam. Amazingly, relative to control, the areas under the concentration time curve and peak plasma concentration of Piracetam were 1.50- and 1.45-fold, respectively, greater in the Morin-pretreated group. However, co-administration of Morin had no significant effect on these parameters. Apart from the aforementioned merits, the results of this study are further confirmed by clinical trials; Piracetam dosages should be adjusted to avoid potential drug interaction when Piracetam is used clinically in combination with Morin and Morin-containing dietary supplements. The in vitro enzyme kinetics were performed to determined km, Vmax & CLins . The in vitro metabolic stability executed for the estimation of metabolic rate constant and half-life of Piracetam. These studies also extrapolate to in vivo intrinsic hepatic clearance (Clint, in vivo ) from in vitro intrinsic hepatic clearance (CLint, in vitro ).

  6. Dose-Dependent Change in Elimination Kinetics of Ethanol due to Shift of Dominant Metabolizing Enzyme from ADH 1 (Class I) to ADH 3 (Class III) in Mouse.

    Science.gov (United States)

    Haseba, Takeshi; Kameyama, Kouji; Mashimo, Keiko; Ohno, Youkichi

    2012-01-01

    ADH 1 and ADH 3 are major two ADH isozymes in the liver, which participate in systemic alcohol metabolism, mainly distributing in parenchymal and in sinusoidal endothelial cells of the liver, respectively. We investigated how these two ADHs contribute to the elimination kinetics of blood ethanol by administering ethanol to mice at various doses, and by measuring liver ADH activity and liver contents of both ADHs. The normalized AUC (AUC/dose) showed a concave increase with an increase in ethanol dose, inversely correlating with β. CL(T) (dose/AUC) linearly correlated with liver ADH activity and also with both the ADH-1 and -3 contents (mg/kg B.W.). When ADH-1 activity was calculated by multiplying ADH-1 content by its V(max⁡)/mg (4.0) and normalized by the ratio of liver ADH activity of each ethanol dose to that of the control, the theoretical ADH-1 activity decreased dose-dependently, correlating with β. On the other hand, the theoretical ADH-3 activity, which was calculated by subtracting ADH-1 activity from liver ADH activity and normalized, increased dose-dependently, correlating with the normalized AUC. These results suggested that the elimination kinetics of blood ethanol in mice was dose-dependently changed, accompanied by a shift of the dominant metabolizing enzyme from ADH 1 to ADH 3.

  7. Dose-Dependent Change in Elimination Kinetics of Ethanol due to Shift of Dominant Metabolizing Enzyme from ADH 1 (Class I to ADH 3 (Class III in Mouse

    Directory of Open Access Journals (Sweden)

    Takeshi Haseba

    2012-01-01

    Full Text Available ADH 1 and ADH 3 are major two ADH isozymes in the liver, which participate in systemic alcohol metabolism, mainly distributing in parenchymal and in sinusoidal endothelial cells of the liver, respectively. We investigated how these two ADHs contribute to the elimination kinetics of blood ethanol by administering ethanol to mice at various doses, and by measuring liver ADH activity and liver contents of both ADHs. The normalized AUC (AUC/dose showed a concave increase with an increase in ethanol dose, inversely correlating with β. CLT (dose/AUC linearly correlated with liver ADH activity and also with both the ADH-1 and -3 contents (mg/kg B.W.. When ADH-1 activity was calculated by multiplying ADH-1 content by its Vmax⁡/mg (4.0 and normalized by the ratio of liver ADH activity of each ethanol dose to that of the control, the theoretical ADH-1 activity decreased dose-dependently, correlating with β. On the other hand, the theoretical ADH-3 activity, which was calculated by subtracting ADH-1 activity from liver ADH activity and normalized, increased dose-dependently, correlating with the normalized AUC. These results suggested that the elimination kinetics of blood ethanol in mice was dose-dependently changed, accompanied by a shift of the dominant metabolizing enzyme from ADH 1 to ADH 3.

  8. Classical galactosaemia revisited

    NARCIS (Netherlands)

    A.M. Bosch

    2006-01-01

    Classical galactosaemia (McKusick 230400) is an: autosomal recessive disorder of galactose metabolism, caused by a deficiency of the enzyme galactose-1-phosphate uridyltransferase (GALT; EC 2.7.712). Most patients present in the neonatal period, after ingestion of galactose, with jaundice, hepatospl

  9. The glucuronidation of R- and S-lorazepam: human liver microsomal kinetics, UDP-glucuronosyltransferase enzyme selectivity, and inhibition by drugs.

    Science.gov (United States)

    Uchaipichat, Verawan; Suthisisang, Chuthamanee; Miners, John O

    2013-06-01

    The widely used hypnosedative-anxiolytic agent R,S-lorazepam is cleared predominantly by conjugation with glucuronic acid in humans, but the enantioselective glucuronidation of lorazepam has received little attention. The present study characterized the kinetics of the separate R and S enantiomers of lorazepam by human liver microsomes (HLMs) and by a panel of recombinant human UDP-glucuronosyltransferase (UGT) enzymes. Respective mean K(m) and V(max) values for R- and S-lorazepam glucuronidation by HLM were 29 ± 8.9 and 36 ± 10 µM, and 7.4 ± 1.9 and 10 ± 3.8 pmol/min ⋅ mg. Microsomal intrinsic clearances were not significantly different, suggesting the in vivo clearances of R- and S-lorazepam are likely to be similar. Both R- and S-lorazepam were glucuronidated by UGT2B4, 2B7, and 2B15, whereas R-lorazepam was additionally metabolized by the extrahepatic enzymes UGT1A7 and 1A10. Based on in vitro clearances and consideration of available in vivo and in vitro data, UGT2B15 is likely to play an important role in the glucuronidation of R- and S-lorazepam. However, the possible contribution of other enzymes and the low activities observed in vitro indicate that the lorazepam enantiomers are of limited use as substrate probes for UGT2B15. To identify potential drug-drug interactions, codeine, fluconazole, ketamine, ketoconazole, methadone, morphine, valproic acid, and zidovudine were screened as inhibitors of R- and S-lorazepam glucuronidation by HLM. In vitro-in vivo extrapolation suggested that, of these drugs, only ketoconazole had the potential to inhibit lorazepam clearance to a clinically significant extent.

  10. Classics Online.

    Science.gov (United States)

    Clayman, Dee L.

    1995-01-01

    Appraises several databases devoted to classical literature. Thesaurus Linguae Graecae (TLG) contains the entire extant corpus of ancient Greek literature, including works on lexicography and historiography, extending into the 15th century. Other works awaiting completion are the Database of Classical Bibliography and a CD-ROM pictorial dictionary…

  11. Classical integrability

    Science.gov (United States)

    Torrielli, Alessandro

    2016-08-01

    We review some essential aspects of classically integrable systems. The detailed outline of the sections consists of: 1. Introduction and motivation, with historical remarks; 2. Liouville theorem and action-angle variables, with examples (harmonic oscillator, Kepler problem); 3. Algebraic tools: Lax pairs, monodromy and transfer matrices, classical r-matrices and exchange relations, non-ultralocal Poisson brackets, with examples (non-linear Schrödinger model, principal chiral field); 4. Features of classical r-matrices: Belavin-Drinfeld theorems, analyticity properties, and lift of the classical structures to quantum groups; 5. Classical inverse scattering method to solve integrable differential equations: soliton solutions, spectral properties and the Gel’fand-Levitan-Marchenko equation, with examples (KdV equation, Sine-Gordon model). Prepared for the Durham Young Researchers Integrability School, organised by the GATIS network. This is part of a collection of lecture notes.

  12. Enzyme efficiency: An open reaction system perspective

    Energy Technology Data Exchange (ETDEWEB)

    Banerjee, Kinshuk, E-mail: kb36@rice.edu [Department of Chemistry, University of Calcutta, Rajabazar Science College Campus, Kolkata 700 009 (India); Bhattacharyya, Kamal, E-mail: pchemkb@gmail.com [Department of Chemistry, University of Calcutta, 92 A.P.C. Road, Kolkata 700 009 (India)

    2015-12-21

    A measure of enzyme efficiency is proposed for an open reaction network that, in suitable form, applies to closed systems as well. The idea originates from the description of classical enzyme kinetics in terms of cycles. We derive analytical expressions for the efficiency measure by treating the network not only deterministically but also stochastically. The latter accounts for any significant amount of noise that can be present in biological systems and hence reveals its impact on efficiency. Numerical verification of the results is also performed. It is found that the deterministic equation overestimates the efficiency, the more so for very small system sizes. Roles of various kinetics parameters and system sizes on the efficiency are thoroughly explored and compared with the standard definition k{sub 2}/K{sub M}. Study of substrate fluctuation also indicates an interesting efficiency-accuracy balance.

  13. Potentiality of yeast Candida sp. SMN04 for degradation of cefdinir, a cephalosporin antibiotic: kinetics, enzyme analysis and biodegradation pathway.

    Science.gov (United States)

    Selvi, A; Das, Devlina; Das, Nilanjana

    2015-01-01

    A new yeast strain isolated from the pharmaceutical wastewater was capable of utilizing cefdinir as a sole carbon source for their growth in mineral medium. The yeast was identified and named as Candida sp. SMN04 based on morphology and 18S-ITS-D1/D2/D3 rRNA sequence analysis. The interaction between factors pH (3.0-9.0), inoculum dosage (1-7%), time (1-11 day) and cefdinir concentration (50-450 mg/L) was studied using a Box-Behnken design. The factors were studied as a result of their effect on cell dry weight (R1; g/L), extended spectrum β-lactamase (ESBL) assay (R2; mm), P450 activity (R3; U/mL) and degradation (R4; %). Maximum values of R1, R2, R3 and R4 were obtained at central values of all the parameters. The isolated yeast strain efficiently degraded 84% of 250 mg L⁻¹ of cefdinir within 6 days with a half-life of 2.97 days and degradation rate constant of 0.2335 per day. Pseudo-first-order model efficiently described the process. Among the various enzymes tested, the order of activity at the end of Day 4 was noted to be: cytochrome P450 (1.76 ± 0.03) > NADPH reductase (1.51 ± 0.20) > manganese peroxidase and amylase (0.66 ± 0.15; 0.66 ± 0.70). Intermediates were successfully characterized by liquid chromatography-mass spectrometry. The opening of the β-lactam ring involving ESBL activity was considered as one of the major steps in the cefdinir degradation process. Fourier transform-infrared spectroscopy analysis showed the absence of spectral vibrations between 1766 and 1519 cm⁻¹ confirming the complete removal of lactam ring during cefdinir degradation. The results of the present study are promising for the use of isolated yeast Candida sp. SMN04 as a potential bioremediation agent.

  14. Unchanged content of oxidative enzymes in fast-twitch muscle fibers and V˙O2 kinetics after intensified training in trained cyclists

    DEFF Research Database (Denmark)

    Christensen, Peter Møller; Gunnarsson, Thomas Gunnar Petursson; Thomassen, Martin;

    2015-01-01

    The present study examined if high intensity training (HIT) could increase the expression of oxidative enzymes in fast-twitch muscle fibers causing a faster oxygen uptake (V˙O2) response during intense (INT), but not moderate (MOD), exercise and reduce the V˙O2 slow component and muscle metabolic...... perturbation during INT. Pulmonary V˙O2 kinetics was determined in eight trained male cyclists (V˙O2-max: 59 ± 4 (means ± SD) mL min(-1) kg(-1)) during MOD (205 ± 12 W ~65% V˙O2-max) and INT (286 ± 17 W ~85% V˙O2-max) exercise before and after a 7-week HIT period (30-sec sprints and 4-min intervals) with a 50...... between MOD and INT. Muscle creatine phosphate was lower (42 ± 15 vs. 66 ± 17 mmol kg DW(-1)) and muscle lactate was higher (40 ± 18 vs. 14 ± 5 mmol kg DW(-1)) at 6 min of INT (P training with a volume reduction did not increase the content...

  15. 竞争性抑制的非稳态酶动力学布尔函数图论研究%Studies on Non-Steady State Enzyme Kinetics of Competitive Inhibition

    Institute of Scientific and Technical Information of China (English)

    赵敏

    2000-01-01

    以非稳态酶动力学的布尔函数图形方法[1],来研究一类竞争性抑制的非稳态酶动力学问题,推导出此类反应的非稳态酶动力学方程,并对此动力学方程进行了讨论,分析了此类竞争性抑制酶反应体系的非稳态酶动力学问题.%This paper is based on the Boolen function graphic analysis in Enzyme kinetics under non-static state and studies a non-static enzyme kinetic dynamic problem of some kind of competitive inhibition. It also deduces the equation of this chemistry and diswsses it. What's more, it analyses the model of non-static enzyme kinetics in the system of such a kind of competitive inhibitive enzyme.

  16. Drug screening in urine by cloned enzyme donor immunoassay (CEDIA) and kinetic interaction of microparticles in solution (KIMS): a comparative study.

    Science.gov (United States)

    Schwettmann, Lutz; Külpmann, Wolf-Rüdiger; Vidal, Christian

    2006-01-01

    Two commercially available drug-screening assays were evaluated: the Roche kinetic interaction of microparticles in solution (KIMS) assay and the Microgenics cloned enzyme donor immunoassay (CEDIA). Urine samples from known drug-abuse patients were analyzed for amphetamines, barbiturates, benzodiazepines, benzoylecgonine, cannabinoids, LSD, methadone and opiates. Samples with discordant findings for the two assays were analyzed by gas chromatography/mass spectrometry (GC/MS) or gas chromatography/electron capture detection (GC/ECD). Amphetamines showed 96.0% concordant results, with two false positive findings by CEDIA, three by KIMS and a further two false negatives by KIMS. Barbiturates showed 99.4% concordant results, with one false negative by KIMS. Benzodiazepines showed 97.4% concordant results, with two false negatives by KIMS (cutoff 100 microg/L, CEDIA cutoff 300 microg/L). Benzoylecgonine showed 17.8% concordant positive and 82.2% concordant negative results and no false finding by either assay. Cannabinoids showed 99.3% concordant results, with one sample negative by KIMS at a cutoff of 50 microg/L and positive by CEDIA (cutoff 25 microg/L). For LSD, 6.7% of findings were not in agreement. Methadone showed 97.5% concordant results, with two false positives by CEDIA, and one false positive and one false negative by KIMS. Opiates showed 96.9% concordant results, with no false KIMS results, but four false positives by CEDIA. The results indicate that the agreement of the CEDIA and KIMS results for the eight drugs is rather good (93.3-100%).

  17. Classical Mechanics

    OpenAIRE

    Gallavotti, Giovanni

    2012-01-01

    This is the English version of a friendly graduate course on Classical Mechanics, containing about 80% of the material I covered during the January-June 1999 semester at IFUG in the Mexican city of Leon. For the Spanish version, see physics/9906066

  18. Modeling chronic hepatitis B or C virus infection during antiviral therapy using an analogy to enzyme kinetics: long-term viral dynamics without rebound and oscillation.

    Science.gov (United States)

    Takayanagi, Toshiaki

    2013-12-01

    The basic model for chronic hepatitis B virus (HBV) or hepatitis C virus (HCV) infection during therapy enables us to analyze short-term viral kinetics. However, the model is not useful for analyzing long-term viral kinetics. Here, I suggest a new model that was obtained by introducing Michaelis-Menten kinetics into the basic model. The new model can exhibit long-term viral kinetics without rebound and oscillation, unlike the basic model. The value of the parameter K in the new model is analogous to the Michaelis constant Km and is predicted to be approximately less than 10(10)/ml.

  19. Research on a novel kinetics experiment of enzyme-catalyzed reaction presented for undergraduate teaching%适用于本科教学的新酶促反应动力学实验研究

    Institute of Scientific and Technical Information of China (English)

    吴梅芬; 王晓岗; 刘亚菲; 许新华

    2015-01-01

    The mechanism of a-chymotrypsin-catalyzed hydrolysis of 4-nitrophenyl trimethylacetate includes the following steps:the reversible enzyme-substrate binding and the enzyme acylation and enzyme deacylation.This mechanism is quite different from the conventional Michaelis-Menten mechanism for enzyme-catalyzed reactions.The corresponding kinetic parameters can be expressed precisely on the basis of chemical kinetics theory,such as Michaelis constant KM ,equilibrium constant K,catalytic rate constant kcat ,and the rate constants of the elementary reactions k 2 and k 3 .The reaction is carried out at 25 °C in a pH = 8.5 buffer of tris(hydroxymethyl)aminomethane (TRIS)by means of spectrophotometry.The kinetic curves are fitted linearly and non-linearly.The kinetic parameters are in agreement with the theoretical prediction.The enzyme product used in this experiment is cheap and the activity is stable. The experimental method is simple enough to be applied in undergraduate physical chemistry laboratory courses.As an ideal model of complex reaction,the experiment will help the students to enhance the understanding and knowledge of chemical kinetics theories.%通过严格的化学反应动力学推导,得到了a-糜蛋白酶催化三甲基乙酸对硝基苯酯水解反应相关的动力学参数的表达式,包括表观米氏常数 KM、酶-底物络合物稳定常数 K 、催化速率常数 kcat 、以及基元反应步骤速率常 k2和 k3等.实验在25℃,pH =8.5的三羟基氨基甲烷缓冲液中进行,用分光光度法测定反应的动力学曲线.数据经过非线性拟合和线性拟合,得到相关的动力学参数,该参数与理论预测一致.该实验酶制剂价格低廉,活性稳定,实验方法简单,可作为本科物理化学实验课程中复杂反应动力学测量的一个很好的实例,加深学生对化学反应动力学相关理论和概念的理解和掌握.

  20. Enzymes involved in the glycidaldehyde (2,3-epoxy-propanal) oxidation step in the kinetic resolution of racemic glycidol (2,3-epoxy-1-propanol) by Acetobacter pasteurianus.

    Science.gov (United States)

    Wandel, U; Machado, S S.; Jongejan, J A.; Duine, J A.

    2001-02-01

    It is already known that kinetic resolution of racemic glycidol (2,3-epoxy-1-propanol) takes place when Acetobacter pasteurianus oxidizes the compound to glycidic acid (2,3-epoxy-propionic acid) with glycidaldehyde (2,3-epoxy-propanal) proposed to be the transient seen in this conversion. Since inhibition affects the feasibility of a process based on this conversion in a negative sense, and the chemical reactivity of glycidaldehyde predicts that it could be the cause for the phenomena observed, it is important to know which enzyme(s) oxidise(s) this compound. To study this, rac.- as well as (R)-glycidaldehyde were prepared by chemical synthesis and analytical methods developed for their determination. It appears that purified quinohemoprotein alcohol dehydrogenase (QH-ADH type II), the enzyme responsible for the kinetic resolution of rac.-glycidol, also catalyses the oxidation of glycidaldehyde. In addition, a preparation exhibiting dye-linked aldehyde dehydrogenase activity for acetaldehyde, most probably originating from molybdohemoprotein aldehyde dehydrogenase (ALDH), which has been described for other Acetic acid bacteria, oxidised glycidaldehyde as well with a preference for the (R)-enantiomer, the selectivity quantified by an enantiomeric ratio (E) value of 7. From a comparison of the apparent kinetic parameter values of QH-ADH and ALDH, it is concluded that ALDH is mainly responsible for the removal of glycidaldehyde in conversions of glycidol catalysed by A. pasteurianus cells. It is shown that the transient observed in rac.-glycidol conversion by whole cells, is indeed (R)-glycidaldehyde. Since both QH-ADH and ALDH are responsible for vinegar production from ethanol by Acetobacters, growth and induction conditions optimal for this process seem also suited to yield cells with high catalytic performance with respect to kinetic resolution of glycidol and prevention of formation of inhibitory concentrations glycidaldehyde.

  1. [Classical taxomomies].

    Science.gov (United States)

    Liubarskiĭ, G Iu

    2006-01-01

    The sequence of classic paradigms in taxonomy that partly replaced each other and partly co-exist is given as follows: the theory of "organ and organism similarity", the naturalistic theory, the descriptive theory, and the phylogenetic theory. The naturalistic classics accepted the notion of "the plan of creation". The rejection of appealing to this plan brought forth certain problems in the formulation of the purpose of taxonomy; these problems were differently solved by the descriptive and the phylogenetic classic traditions. The difficulties of the current paradigms arising from the loss of a "strong purpose", a problem to be solved by taxonomists that is to be clear and interesting to a wide range of non-professionals. The paradox of formalization led to the losing of content of the methods due to their formalization. To attract attention to taxonomy, a new "image of the results" of its work that would be interesting to the non-professionals is necessary. The co-existence of different methods of reseach applied to different groups of facts leads to the loss of integrity of the research. It is not only that the taxon becomes a hypothesis and such hypotheses multiply. The comparison of these hypotheses is problematic, because each of them is supported by its own independent scope of facts. Because of the existence of a fundamental meronotaxonomic discrepancy, taxonomic systems based on different groups of characters appear to be incomparable, being rather systems of characters than systems of taxa. Systems of characters are not directly comparable with each other; they can be compared only through appealing to taxa, but taxa themselves exist only in the form of a number of hypotheses. Consequently, each separate taxonomic approach creates its own nature, its own subject of research. Therefore, it is necessary to describe the subject of research correctly (and indicate the purpose of research), as well as to distinguish clearly between results achieved through

  2. Alkylating enzymes.

    Science.gov (United States)

    Wessjohann, Ludger A; Keim, Jeanette; Weigel, Benjamin; Dippe, Martin

    2013-04-01

    Chemospecific and regiospecific modifications of natural products by methyl, prenyl, or C-glycosyl moieties are a challenging and cumbersome task in organic synthesis. Because of the availability of an increasing number of stable and selective transferases and cofactor regeneration processes, enzyme-assisted strategies turn out to be promising alternatives to classical synthesis. Two categories of alkylating enzymes become increasingly relevant for applications: firstly prenyltransferases and terpene synthases (including terpene cyclases), which are used in the production of terpenoids such as artemisinin, or meroterpenoids like alkylated phenolics and indoles, and secondly methyltransferases, which modify flavonoids and alkaloids to yield products with a specific methylation pattern such as 7-O-methylaromadendrin and scopolamine.

  3. Evaluation of an Erns-based enzyme-linked immunosorbent assay to distinguish Classical swine fever virus-infected pigs from pigs vaccinated with CP7_E2alf.

    Science.gov (United States)

    Pannhorst, Katrin; Fröhlich, Andreas; Staubach, Christoph; Meyer, Denise; Blome, Sandra; Becher, Paul

    2015-07-01

    Infections with Classical swine fever virus (CSFV) are a major economic threat to pig production. To combat CSF outbreaks and to maintain trade, new marker vaccines were developed that allow differentiation of infected from vaccinated animals (DIVA principle). The chimeric pestivirus CP7_E2alf was shown to be safe and efficacious. Its DIVA strategy is based on the detection of CSFV E(rns)-specific antibodies that are only developed on infection. However, for the new marker vaccine to be considered a valuable control tool, a validated discriminatory assay is needed. One promising candidate is the already commercially available enzyme-linked immunosorbent assay, PrioCHECK CSFV E(rns) ELISA (Prionics BV, Lelystad, The Netherlands). Four laboratories of different European Union member states tested 530 serum samples and country-specific field sera from domestic pigs and wild boar. The ELISA displayed a good robustness. However, based on its reproducibility and repeatability, ranges rather than single values for diagnostic sensitivity and specificity were defined. The ELISA displayed a sensitivity of 90-98% with sera from CSFV-infected domestic pigs. A specificity of 89-96% was calculated with sera from domestic pigs vaccinated once with CP7_E2alf. The ELISA detected CSFV infections in vaccinated domestic pigs with a sensitivity of 82-94%. The sensitivity was lower with sera taken ≤21 days post-challenge indicating that the stage of CSFV infection had a considerable influence on testing. Taken together, the PrioCHECK CSFV E(rns) ELISA can be used for detection of CSFV infections in CP7_E2alf-vaccinated and nonvaccinated domestic pig populations, but should only be applied on a herd basis by testing a defined number of animals.

  4. Adenine phosphoribosyltransferase from Sulfolobus solfataricus is an enzyme with unusual kinetic properties and a crystal structure that suggests it evolved from a 6-oxopurine phosphoribosyltransferase

    DEFF Research Database (Denmark)

    Jensen, Kaj Frank; Hansen, Michael Riis; Jensen, Kristine Steen

    2015-01-01

    The adenine phosphoribosyltransferase (APRTase) encoded by the open reading frame SSO2342 of Sulfolobus solfataricus P2, was subjected to crystallographic, kinetic and ligand binding analyses. The enzyme forms dimers in solution and in the crystals, and binds one molecule of the reactants 5......-phosphoribosyl-α-1-pyrophosphate (PRPP) and adenine or the product AMP or the inhibitor ADP in each active site. The individual subunit adopts an overall structure that resembles a 6-oxopurine phosphoribosyltransferase (PRTase) more than known APRTases implying that APRT functionality in Crenarchaeotae has its...... undergoes a conformational change upon binding of adenine and phosphate resulting in a slight contraction of the active site. The inhibitor, ADP binds like the product AMP with both the α- and β-phosphates occupying the 5’-phosphoribosyl binding site. The enzyme shows activity over a wide p...

  5. A chimeric LysK-lysostaphin fusion enzyme lysing Staphylococcus aureus cells: a study of both kinetics of inactivation and specifics of interaction with anionic polymers

    Science.gov (United States)

    A staphylolytic fusion protein (K-L) was created, harboring three unique lytic activities comprised of the LysK CHAP endopeptidase, and amidase domains, and the lysostaphin glycyl-glycine endopeptidase domain. To assess the potential of possible therapeutic applications, the kinetic behavior of K-L...

  6. A Digital Microfluidic System for the Investigation of Pre-Steady-State Enzyme Kinetics Using Rapid Quenching with MALDI-TOF Mass Spectrometry

    NARCIS (Netherlands)

    Nichols, Kevin Paul; Gardeniers, Han J.G.E.

    2007-01-01

    A digital microfluidic system based on electrowetting has been developed to facilitate the investigation of pre-steady-state reaction kinetics using rapid quenching and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The device consists of individually ad

  7. King-Altman作图法对酶可逆抑制反应动力学的应用%Application of King-Altman Plot in the Kinetics for the Reversibly Inhibited Enzyme-catalyzed Reaction

    Institute of Scientific and Technical Information of China (English)

    杨明莉; 任建敏

    2001-01-01

      With the King-Altman plot, the kinetics equation of a single substrate enzyme-catalyzed reaction inhibited by a reversible inhibitor was attained. With the help of the general kinetics equation from an enzyme-catalyzed reaction obeying partially reversible inhibiting mechanism, the reaction mechanisms and the characteristics for partially and fully competitive inhibition, partially and fully noncompetitive inhibition, partially and fully uncompetitive inhibition were discussed.%  本文用King-Altman作图法推导了可逆抑制酶促单底物反应动力学方程,并借助于部分可逆抑制单底物酶促反应动力学方程对完全与部分的竞争性抑制作用(PC与FC)、完全与部分非竞争性抑制作用(FNC与PNC)、完全与部分反竞争性抑制作用(FUC与PUC)和完全与部分混合抑制作用(FM与PM)的动力学机理与特点进行了讨论。

  8. Partial purification, characterization, and kinetic studies of a low-molecular-weight, alkali-tolerant chitinase enzyme from Bacillus subtilis JN032305, A potential biocontrol strain.

    Science.gov (United States)

    Shivakumar, Srividya; Karmali, Anika Nayak; Ruhimbana, Charles

    2014-01-01

    A new alkalophilic low-molecular-mass chitinase of 14 kD from the potent biocontrol agent Bacillus subtilis JN032305 was partially purified and enzymology of the chitinase was studied. The enzyme showed optimal pH of 9.0 and temperature of 50°C. The enzyme was found stable during the 60-min incubation at 50 °C. The chitinase was inhibited by group specific agents like IAA, DAN, TLCK, and SDS and metal ions Mg(2+), Ca(2+), Fe(2+), Mn(2+), Ba(2+), and Hg(2+), whereas Zn(2+) did not show significant inhibitory effect against the chitinase. PMSF partially inhibited the enzyme. Substrates specificity tests indicated that the enzyme showed 75% of relative activity on glycol chitin, 58% on carboxymethylcellulose (CMC), 33% on chitin flakes, and 166% laminarin compared to that on colloidal chitin. The enzyme also hydrolyzed 4-methylumbelliferyl-N-acetyl-D-glucosaminide, indicating its chitobiase activity. The chitinase of this study has broad specificity, which could hydrolyze not only the glycosidic bond in GlcNAc-GlcNAc but also that of related carbohydrates with glycosidic linkages. The partially purified chitinase not only showed antifungal activity against Rhizoctonia solani and Colletotrichum gloeosporioides, two potent phytopathogens of chilli, but also increased the germination of chilli seeds when infected with the two potent phytopathogenic fungi.

  9. Reassessment of the transhydrogenase/malate shunt pathway in Clostridium thermocellum ATCC 27405 through kinetic characterization of malic enzyme and malate dehydrogenase.

    Science.gov (United States)

    Taillefer, M; Rydzak, T; Levin, D B; Oresnik, I J; Sparling, R

    2015-04-01

    Clostridium thermocellum produces ethanol as one of its major end products from direct fermentation of cellulosic biomass. Therefore, it is viewed as an attractive model for the production of biofuels via consolidated bioprocessing. However, a better understanding of the metabolic pathways, along with their putative regulation, could lead to improved strategies for increasing the production of ethanol. In the absence of an annotated pyruvate kinase in the genome, alternate means of generating pyruvate have been sought. Previous proteomic and transcriptomic work detected high levels of a malate dehydrogenase and malic enzyme, which may be used as part of a malate shunt for the generation of pyruvate from phosphoenolpyruvate. The purification and characterization of the malate dehydrogenase and malic enzyme are described in order to elucidate their putative roles in malate shunt and their potential role in C. thermocellum metabolism. The malate dehydrogenase catalyzed the reduction of oxaloacetate to malate utilizing NADH or NADPH with a kcat of 45.8 s(-1) or 14.9 s(-1), respectively, resulting in a 12-fold increase in catalytic efficiency when using NADH over NADPH. The malic enzyme displayed reversible malate decarboxylation activity with a kcat of 520.8 s(-1). The malic enzyme used NADP(+) as a cofactor along with NH4 (+) and Mn(2+) as activators. Pyrophosphate was found to be a potent inhibitor of malic enzyme activity, with a Ki of 0.036 mM. We propose a putative regulatory mechanism of the malate shunt by pyrophosphate and NH4 (+) based on the characterization of the malate dehydrogenase and malic enzyme.

  10. Forward design of a complex enzyme cascade reaction

    Science.gov (United States)

    Hold, Christoph; Billerbeck, Sonja; Panke, Sven

    2016-01-01

    Enzymatic reaction networks are unique in that one can operate a large number of reactions under the same set of conditions concomitantly in one pot, but the nonlinear kinetics of the enzymes and the resulting system complexity have so far defeated rational design processes for the construction of such complex cascade reactions. Here we demonstrate the forward design of an in vitro 10-membered system using enzymes from highly regulated biological processes such as glycolysis. For this, we adapt the characterization of the biochemical system to the needs of classical engineering systems theory: we combine online mass spectrometry and continuous system operation to apply standard system theory input functions and to use the detailed dynamic system responses to parameterize a model of sufficient quality for forward design. This allows the facile optimization of a 10-enzyme cascade reaction for fine chemical production purposes. PMID:27677244

  11. The Reaction Kinetics of 3-Hydroxybenzoate 6-Hydroxylase from Rhodococcus jostii RHA1 Provide an Understanding of the para-Hydroxylation Enzyme Catalytic Cycle

    NARCIS (Netherlands)

    Sucharitakul, J.; Tongsook, C.; Pakotiprapha, D.; Berkel, van W.J.H.; Chaiyen, P.

    2013-01-01

    3-Hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1 is an NADH-specific flavoprotein monooxygenase that catalyzes the para-hydroxylation of 3-hydroxybenzoate (3HB) to form 2,5-dihydroxybenzoate (2,5-DHB). Based on results from stopped-flow spectrophotometry, the reduced enzyme-3HB c

  12. Unchanged content of oxidative enzymes in fast-twitch muscle fibers and V˙O2 kinetics after intensified training in trained cyclists

    DEFF Research Database (Denmark)

    Christensen, Peter Møller; Gunnarsson, Thomas Gunnar Petursson; Thomassen, Martin;

    2015-01-01

    DW(-1) min(-1)) of CS (56 ± 8 post-HIT vs. 59 ± 10 pre-HIT), HAD (27 ± 6 vs. 29 ± 3) and PFK (340 ± 69 vs. 318 ± 105) and the capillary to fiber ratio (2.30 ± 0.16 vs. 2.38 ± 0.20) was unaltered following HIT. V˙O2 kinetics was unchanged with HIT and the speed of the primary response did not differ...

  13. Deuterium kinetic isotope effects in heterotetrameric sarcosine oxidase from Corynebacterium sp. U-96: the anionic form of the substrate in the enzyme-substrate complex is a reactive species.

    Science.gov (United States)

    Saito, Mutsumi; Itoh, Ai; Suzuki, Haruo

    2012-06-01

    Heterotetrameric sarcosine oxidase is a flavoprotein that catalyses the oxidative demethylation of sarcosine. It is thought that the dehydrogenated substrate is the anionic form of sarcosine. To verify this assumption, the rate of flavin-adenine dinucleotide (FAD) reduction (k(red)) was analysed using protiated and deuterated sarcosine (N-methyl-d(3)-Gly) at various pH values using stopped-flow method. By increasing the pH from 6.2 to 9.8, k(red) increased for both substrates and reached a plateau, but the pK(a) value (reflecting the ionization of the enzyme-substrate complex) was 6.8 and 7.1 for protiated and deuterated sarcosine, respectively, and the kinetic isotope effect of k(red) decreased from approximately 19 to 8, indicating deprotonation of the bound sarcosine. The k(red)/K(d) (K(d), sarcosine dissociation constant) increased with increasing pH and reached a plateau. The pK (reflecting the ionization of free enzyme or free sarcosine) was 7.0 for both substrates, suggesting deprotonation of the βLys358 residue, which has a pK(a) of 6.7, as the pK(a) of the free sarcosine amine proton was determined to be approximately 10.1. These results indicate that the amine proton of sarcosine is transferred to the unprotonated Lys residue in the enzyme-substrate complex.

  14. Classical and quantum free motions in the tomographic probability representation

    CERN Document Server

    Man'ko, Vladimir I

    2011-01-01

    Based on a geometric picture, the example of free particle motion for both classical and quantum domains is considered in the tomographic probability representation. Wave functions and density operators as well as optical and symplectic tomograms are obtained as solutions of kinetic classical and quantum equations for the state tomograms. The difference of tomograms of free particle for classical and quantum states is discussed.

  15. 蛇床夫内酯在大鼠肝微粒体中的酶促反应动力学研究%Enzyme Kinetics of Cnidilin Metabolism in Rat Liver Microsomes

    Institute of Scientific and Technical Information of China (English)

    霍好利; 苑霖; 刘西哲; 曹亮; 常路; 张兰桐

    2011-01-01

    OBJECTIVE To study the enzyme kinetics of cnidilin metabolism in rat liver microsomes. METHODS A LC-MS method was established for quantitative analysis of two metabolites, 3", 8-methoxy-isoimperatorin ( Ml ) and 5"-hydroxyl-8-methoxy-iso-imperatorin (M2) , with pimpinellin as internal standard. Lineweave-Burk graphic method was used to calculate the enzyme kinetic parameters, maximum reaction rate ( Vmn ) and Km. RESULTS The enzyme kinetics parameters of Ml were as follows; Kmax = 1. 152 5 jimol ? (min ? Mg pro) "' , and Km = 133. 156 6 n,mol ? L~'. And the parameters of M2 were: Vma = 1. 630 8 ujnol ? ( min ? Mg pro) "' , and Km =292. 857 1 jjunol ? L '. CONCLUSION The method is simple, specific and reliable, which is suitable for the in vitro metabolism research of cnidilin.%目的 研究蛇床夫内酯在大鼠肝微粒体中的酶促反应动力学.方法 以回芹内酯为内标,采用液质联用(LC/MS)的方法测定蛇床夫内酯在体外代谢中的代谢产物3″,8-甲氧基异欧前胡内酯(M1)和5″-羟基-8-甲氧基异欧前胡内酯(M2).用Linewrave-Burk作图分析数据,计算酶促动力学常数.结果 代谢物M1的Vmax=1.152 5μmol·(min·mg pro) -,Km=133.156 6 μmol·L-1,M2的Vmax=1.630 8 μmol·(min·mg pro)-1,Km=292.8571 μmol·L-1.结论 该方法简单、快速、可靠,适用于蛇床夫内酯的体外代谢研究.

  16. Purification and immobilization of L-arginase from thermotolerant Penicillium chrysogenum KJ185377.1; with unique kinetic properties as thermostable anticancer enzyme.

    Science.gov (United States)

    El-Sayed, Ashraf S; Shindia, Ahmed A; Diab, Ayman A; Rady, Amgad M

    2014-10-18

    L-Arginase, hydrolyzing L-arginine to L-ornithine and urea, is a powerful anticancer, L-arginine-depleting agent, against argininosuccinate synthase expressing tumors. Otherwise, the higher antigenicity and lower thermal stability of this enzyme was the main biochemical hurdles. Since, the intrinsic thermal stability of enzymes follow the physiological temperature of their producer, thus, characterization of L-arginase from thermotolerant Penicillium chrysogenum was the objective of this study. L-Arginase (Arg) was purified to its homogeneity from P. chrysogenum by 10.1-fold, with 37.0 kDa under denaturing PAGE, optimum reaction at 50 °C, pH stability (6.8-7.9), with highest molar ratio of constitutional arginine, glutamic acid, lysine and aspartic acid. The purified enzyme was PEGylated and immobilized on chitosan, with 41.9 and 22.1 % yield of immobilization. At 40 °C, the T1/2 value of free-Arg, PEG-Arg and Chit-Arg was 10.4, 15.6, 20.5 h, respectively. The free-Arg and Chit-Arg have a higher affinity to L-arginine (K m 4.8 mM), while, PEG-Arg affinity was decreased by about 3 fold (K m 15.2 mM). The inhibitory constants to the free and PEG-Arg were relatively similar towards HA and PPG. The IC50 for the free enzyme against HEPG-2 and A549 tumor cells was 0.136 and 0.165 U/ml, comparing to 0.232 and 0.496 U/ml for PEG-Arg, respectively. The in vivo T1/2 to the free Arg and PEG-Arg was 16.4 and 20.4 h, respectively as holo-enzyme. The residual L-arginine level upon using free Arg was 156.9 and 144.5 µM, after 6 and 8 h, respectively, regarding to initials at 253.6 µM, while for Peg-Arg the level of L-arginine was nil till 7 h of initial dosing. The titer of IgG was induced by 10-15 % in response to free-Arg after 28 days comparing to IgG titer for PEG-Arg.

  17. Multiple alternative substrate kinetics.

    Science.gov (United States)

    Anderson, Vernon E

    2015-11-01

    The specificity of enzymes for their respective substrates has been a focal point of enzyme kinetics since the initial characterization of metabolic chemistry. Various processes to quantify an enzyme's specificity using kinetics have been utilized over the decades. Fersht's definition of the ratio kcat/Km for two different substrates as the "specificity constant" (ref [7]), based on the premise that the important specificity existed when the substrates were competing in the same reaction, has become a consensus standard for enzymes obeying Michaelis-Menten kinetics. The expansion of the theory for the determination of the relative specificity constants for a very large number of competing substrates, e.g. those present in a combinatorial library, in a single reaction mixture has been developed in this contribution. The ratio of kcat/Km for isotopologs has also become a standard in mechanistic enzymology where kinetic isotope effects have been measured by the development of internal competition experiments with extreme precision. This contribution extends the theory of kinetic isotope effects to internal competition between three isotopologs present at non-tracer concentrations in the same reaction mix. This article is part of a special issue titled: Enzyme Transition States from Theory and Experiment.

  18. Studies on Non-steady Enzyme Kinetics of Competitive Inhibition of Bienzyme System%二酶系统竞争性抑制的非稳态动力学

    Institute of Scientific and Technical Information of China (English)

    赵敏

    2001-01-01

    The Boolean function graphic method is applied to the analysis of non-steady state enzyme kinetics of competitive inhibition of bienzyme system.The rate equation of the bienzyme reaction system is discussed.%: 以非稳态酶动力学的布尔函数图形方法,研究了一类二酶系统竞争性抑制的非稳态动力学,推导出此类反应的非稳态酶动力学方程,并对其进行了讨论,分析了这类二酶系统竞争性抑制的非稳态动力学过程。

  19. Effects of cellulase and xylanase enzymes mixed with increasing doses ofSalix babylonicaextract onin vitrorumengas production kinetics of a mixture of corn silage with concentrate

    Institute of Scientific and Technical Information of China (English)

    Abdelfattah Z M Salem; German Buenda-Rodrguez; Mona M M Elghandour; Mara A Mariezcurrena Berasain; Francisco J Pea Jimnez; Alberto B Pliego; Juan C V Chagoyn; Mara A Cerrillo; Miguel A Rodrguez

    2015-01-01

    Anin vitro gas production (GP) technique was used to investigate the effects of combining different doses ofSalix babylonicaextract (SB) with exogenous ifbrolytic enzymes (EZ) based on xylanase (X) and celulase (C), or their mixture (XC; 1:1 v/v) onin vitrofermentation characteristics of a total mixed ration of corn silage and concentrate mixture (50:50, w/w)as substrate. Four levels of SB (0, 0.6, 1.2 and 1.8 mL g–1 dry matter (DM))andfour supplemental styles of EZ (1 µL g–1 DM; control (no enzymes), X, C and XC (1:1, v/v) were used in a 4×4 factorial arrangement.In vitro GP (mL g–1 DM) were recorded at 2, 4, 6, 8, 10, 12, 24, 36, 48 and 72 h of incubation. After 72 h, the incubation process was stopped and supernatant pH was determined, and then ifltered to determine dry matter degradability (DMD). Fermentation parameters, such as the 24 h gas yield (GY24),in vitro organic matter digestibility (OMD), metabolizable energy (ME), short chain fatty acid concentrations (SCFA), and microbial crude protein production (MCP) were also estimated. Results indicated that there was a SB´EZ interaction (P0.05) on OMD, pH, ME, GY24, SCFA and MP. The combination of SB with EZ increased (P<0.001) OMD, ME, SCFA, PF72 and GP24, whereas there was no impact on pH. It could be concluded that addition of SB extract, C, and X effectively improved thein vitro rumen fermentation, and the combination of enzyme with SB extract at the level of 1.2 mL g–1 was more effective than the other treatments.

  20. Peroxisomal multifunctional enzyme type 2 from the fruitfly: dehydrogenase and hydratase act as separate entities, as revealed by structure and kinetics.

    Science.gov (United States)

    Haataja, Tatu J K; Koski, M Kristian; Hiltunen, J Kalervo; Glumoff, Tuomo

    2011-05-01

    All of the peroxisomal β-oxidation pathways characterized thus far house at least one MFE (multifunctional enzyme) catalysing two out of four reactions of the spiral. MFE type 2 proteins from various species display great variation in domain composition and predicted substrate preference. The gene CG3415 encodes for Drosophila melanogaster MFE-2 (DmMFE-2), complements the Saccharomyces cerevisiae MFE-2 deletion strain, and the recombinant protein displays both MFE-2 enzymatic activities in vitro. The resolved crystal structure is the first one for a full-length MFE-2 revealing the assembly of domains, and the data can also be transferred to structure-function studies for other MFE-2 proteins. The structure explains the necessity of dimerization. The lack of substrate channelling is proposed based on both the structural features, as well as by the fact that hydration and dehydrogenation activities of MFE-2, if produced as separate enzymes, are equally efficient in catalysis as the full-length MFE-2.

  1. Characterization of the Enantioselective Properties of the Quinohemoprotein Alcohol Dehydrogenase of Acetobacter pasteurianus LMG 1635. 1. Different Enantiomeric Ratios of Whole Cells and Purified Enzyme in the Kinetic Resolution of Racemic Glycidol.

    Science.gov (United States)

    Machado, S S; Wandel, U; Jongejan, J A; Straathof, A J; Duine, J A

    1999-01-01

    Resting cells of Acetobacter pasteurianus LMG 1635 (ATCC 12874) show appreciable enantioselectivity (E=16-18) in the oxidative kinetic resolution of racemic 2,3-epoxy-1-propanol, glycidol. Distinctly lower values (E=7-9) are observed for the ferricyanide-coupled oxidation of glycidol by the isolated quinohemoprotein alcohol dehydrogenase, QH-ADH, which is responsible for the enantiospecific oxidation step in whole cells. The accuracy of E-values from conversion experiments could be verified using complementary methods for the measurement of enantiomeric ratios. Effects of pH, detergent, the use of artificial electron acceptors, and the presence of intermediate aldehydes, could be accounted for. Measurements of E-values at successive stages of the purification showed that the drop in enantioselectivity correlates with the separation of QH-ADH from the cytoplasmic membrane. It is argued that the native arrangement of QH-ADH in the membrane-associated complex favors the higher E-values. The consequences of these findings for the use of whole cells versus purified enzymes in biocatalytic kinetic resolutions of chiral alcohols are discussed.

  2. Enzyme Teaching by a Virtual Laboratory

    Directory of Open Access Journals (Sweden)

    J.K. Sugai

    2010-05-01

    Full Text Available Biochemistry learning demands skills to obtaining and interpreting the experimental data. In a classical model of teaching involve student’s hands-on participation. However this model is expensive, not safe and should be carried out in a short and limited time course. With utilization of educational software these disadvantages are overcome, since the virtual activity could be realized at free full access, and is a tool for individual study. The aim of the present work is to present educational software focused on a virtual for undergraduate student of biochemistry courses. The software development was performed with the help of concept maps, ISIS Draw, ADOBE Photoshop and FLASH MX Program applied on the subject salivary amylase. It was possible to present the basic methodologies for study of the kinetic of enzyme. The substrate (starch consumption was determinate by iodine reaction, while the products (reducing sugars formation was evaluated by cupper-alkaline reaction. The protocols of the virtual experiments are present verbally as well as a subtitle. A set of exercises are disposable, which allowed an auto evaluation and a review of the subject. The experimental treatment involved the presentation of this hypermedia for Nutrition and Dentistry/UFSC undergraduate students as a tool for better comprehension of the theme and promoted the understanding of the kinetic of enzyme.

  3. Inhibition of cytochrome P450 enzymes by saturated and unsaturated fatty acids in human liver microsomes, characterization of enzyme kinetics in the presence of bovine serum albumin (0.1 and 1.0% w/v) and in vitro - in vivo extrapolation of hepatic clearance.

    Science.gov (United States)

    Palacharla, Raghava Choudary; Uthukam, Venkatesham; Manoharan, Arunkumar; Ponnamaneni, Ranjith Kumar; Padala, Nagasurya Prakash; Boggavarapu, Rajesh Kumar; Bhyrapuneni, Gopinadh; Ajjala, Devender Reddy; Nirogi, Ramakrishna

    2017-04-01

    The objective of the study was to determine the effect of fatty acids on CYP enzymes and the effect of BSA on intrinsic clearance of probe substrates. The inhibitory effect of thirteen fatty acids including saturated, mono-unsaturated and polyunsaturated fatty acids on CYP enzymes, kinetic parameters and intrinsic clearance values of nine CYP marker probe substrate reactions in the absence and presence of BSA (0.1 and 1.0% w/v) were characterized in human liver microsomes. The results demonstrate that most of the unsaturated fatty acids showed marked inhibition towards CYP2C8 mediated amodiaquine N-deethylation followed by inhibition of CYP2C9 and CYP2B6 mediated activities. The addition of 0.1% BSA in the incubation markedly improved the unbound intrinsic clearance values of probe substrates by reducing the Km values with little or no effect on maximal velocity. The addition of BSA (0.1 and 1.0% w/v) did not influence the unbound intrinsic clearance of marker reactions for CYP2A6, and CYP3A4 enzymes. The addition of 0.1% w/v BSA is sufficient to determine the intrinsic clearance of marker probe reactions by metabolite formation approach. The predicted hepatic clearance values for the substrates using the well-stirred model, in the presence of BSA (0.1% BSA), are comparable to the in vivo hepatic clearance values.

  4. Study on Molecular Cloning and Enzyme Kinetics of Anabaena α-Phycocyanin Lyase%藻蓝蛋白α亚基裂合酶的分子克隆和酶动力学研究

    Institute of Scientific and Technical Information of China (English)

    苏平; 周明

    2012-01-01

    To comparatively study the differences on structures and functions of phycocyanin lyase CpcE/F from different cyanobacteria, CpcE/F from Anabaena sp. PCC 7120 were cloned and expressed greatly. The reconstitution of PCB and CpcA from Mastigocladus laminosus PCC 7603 in vitro using the overexpressed lyase Anabaena sp. PCC 7120 CpcE/F showed that CpcE/F from Anabaena sp. PCC 7120 is the specific enzyme for biosynthesis of α-PC, furthermore, the enzyme kinetics of PcE/F lyase were studied preliminarily.%为了比较研究不同藻种中藻蓝蛋白裂合酶CpcE/F的结构与功能的差异,对Anabaena sp.PCC 7120中的CpcE/F进行克隆,并进行大量表达,将表达的裂合酶CpcE/F用于藻蓝胆素(PCB)与Mastigocladus laminosus PCC 7603藻蓝蛋白α-亚基(α-PC)脱辅基蛋白(CpcA)的体外重组,得到天然活性的α-PC,从而表明CpcE/F所编码的蛋白质是α-PC生物合成的裂合酶,并对CpcE/F的酶动力学进行了初步研究.

  5. What classicality? Decoherence and Bohr's classical concepts

    CERN Document Server

    Schlosshauer, Maximilian

    2010-01-01

    Niels Bohr famously insisted on the indispensability of what he termed "classical concepts." In the context of the decoherence program, on the other hand, it has become fashionable to talk about the "dynamical emergence of classicality" from the quantum formalism alone. Does this mean that decoherence challenges Bohr's dictum and signifies a break with the Copenhagen interpretation-for example, that classical concepts do not need to be assumed but can be derived? In this paper we'll try to shine some light down the murky waters where formalism and philosophy cohabitate. To begin, we'll clarify the notion of classicality in the decoherence description. We'll then discuss Bohr's and Heisenberg's take on the quantum-classical problem and reflect on different meanings of the terms "classicality" and "classical concepts" in the writings of Bohr and his followers. This analysis will allow us to put forward some tentative suggestions for how we may better understand the relation between decoherence-induced classical...

  6. Enzyme Therapy in Non-classic Pompe’s Disease: Safety and efficacy of recombinant human a-glucosidase from milk of transgenic rabbits and from Chinese hamster ovary cells

    NARCIS (Netherlands)

    L.P.F. Winkel (Léon)

    2004-01-01

    textabstractPompe’s disease is an inherited metabolic illness, caused by an inherited deficiency of an enzyme, called acid a-glucosidase. Acid a-glucosidase is a protein that breaks down glycogen (a chain of glucose molecules) into glucose (a single molecule), in a specific compartment of the cel

  7. Pectic enzymes

    NARCIS (Netherlands)

    Benen, J.A.E.; Voragen, A.G.J.; Visser, J.

    2003-01-01

    The pectic enzymes comprise a diverse group of enzymes. They consist of main-chain depolymerases and esterases active on methyl- and acetylesters of galacturonosyl uronic acid residues. The depolymerizing enzymes comprise hydrolases as wel as lyases

  8. Enzyme assays.

    Science.gov (United States)

    Reymond, Jean-Louis; Fluxà, Viviana S; Maillard, Noélie

    2009-01-07

    Enzyme assays are analytical tools to visualize enzyme activities. In recent years a large variety of enzyme assays have been developed to assist the discovery and optimization of industrial enzymes, in particular for "white biotechnology" where selective enzymes are used with great success for economically viable, mild and environmentally benign production processes. The present article highlights the aspects of fluorogenic and chromogenic substrates, sensors, and enzyme fingerprinting, which are our particular areas of interest.

  9. 青霉素酰化酶交联酶聚体的制备及热失活动力学%Preparation and Thermal Kinetic Deactivation of Cross-linked Enzyme Aggregates of Penicillin Acylase

    Institute of Scientific and Technical Information of China (English)

    慕洋洋; 甄倩楠; 王梦凡; 齐崴; 苏荣欣; 何志敏

    2014-01-01

    以0.53 g/ mL 硫酸铵为沉淀剂,0.35%(体积分数)戊二醛为交联剂制得青霉素酰化酶交联酶聚体(CLEAs),酶活收率30.1%,其最适温度(57℃)比游离酶提高10℃,最适 pH(10.0)向碱性偏移1.7个单位.对比游离酶及其 CLEAs 的热稳定性和热失活动力学模型发现,游离青霉素酰化酶制成 CLEAs 后,其热失活动力学模型由一步失活转变为连串失活,失活反应活化能由248.8 kJ/ mol 增加至549.2 kJ/ mol,对CLEAs 热稳定性大幅提高的原因进行了解释. CLEAs 重复利用7次后,酶活保留56%以上,具有良好的重复利用性.%Cross-linked enzyme aggregates(CLEAs) is an efficient approach to obtain immobilized enzymes without the use of any pre-existing carriers. The preparation of CLEAs usually consisted of two simple steps:precipitation and cross-linking. In this work, CLEAs of penicillin acylase was preparaed using 50% ammo-nium sulphate solution as precipitant and 0. 35% glutaraldehyde as cross-linking agent. The resulting CLEAs obtained an activity yield of 30. 1% and higher optimal temperature (57 ℃) than free penicillin acylase (47 ℃) as well as an obvious shift of optimal pH from 8. 3 to 10. 0. Moreover, compared with free enzyme, the thermal stability of CLEAs was largely enhanced which facilitated the application of penicillin acylase in high-temperature reaction systems. Based on the thermal deactivation kinetics study, it was found that the deactivation model of penicillin acylase changed from one-step model to serial model through immobilizing as CLEAs. The higher deactivation energy of CLEAs(549. 2 kJ/ mol) than free enzyme(248. 8 kJ/ mol) also ex-plained the excellent stability of CLEAs under high temperature environment. In addition, CLEAs exhibited favorable reusability after using 7 times.

  10. Molecular dynamics simulations of classical stopping power.

    Science.gov (United States)

    Grabowski, Paul E; Surh, Michael P; Richards, David F; Graziani, Frank R; Murillo, Michael S

    2013-11-22

    Molecular dynamics can provide very accurate tests of classical kinetic theory; for example, unambiguous comparisons can be made for classical particles interacting via a repulsive 1/r potential. The plasma stopping power problem, of great interest in its own right, provides an especially stringent test of a velocity-dependent transport property. We have performed large-scale (~10(4)-10(6) particles) molecular dynamics simulations of charged-particle stopping in a classical electron gas that span the weak to moderately strong intratarget coupling regimes. Projectile-target coupling is varied with projectile charge and velocity. Comparisons are made with disparate kinetic theories (both Boltzmann and Lenard-Balescu classes) and fully convergent theories to establish regimes of validity. We extend these various stopping models to improve agreement with the MD data and provide a useful fit to our results.

  11. Enzyme Kinetics of Norisoboldine Glucuronidation in Rat Liver Microsomes%去甲异波尔定在大鼠肝微粒体中葡萄糖醛酸化代谢动力学研究

    Institute of Scientific and Technical Information of China (English)

    陈建忠; 侴桂新; 王长虹; 王峥涛

    2012-01-01

    Objective: To investigate the enzyme kinetics of norisoboldine glucuronidation in rat liver microsomes. Method: An UPLC-MS method was developed for determination of norisoboldine-9-O-α-glucuronide, a major metabolite of norisoboldine, in liver microsomes incubation system with sinomenine as internal standard. An optimum incubation system was found and the enzyme kinetics parameters of norisoboldine-9-O-α-glucuronide were analyzed and calculated with Linewearve-Burk graphic method. Result: The Km Vmax and CLin1 ( Vmax/Ka) of norisoboldine-9-O-α-glucuronide were 40. 7 μmol·L-1 , 909. 1 pmol·(min·mg pro)-1 and 22.3 μL·min-1·mg -1, respectively. Conclusion: This method is simple, specific and reliable, which is suitable for the in vitro research of norisoboldine glucuronidation.%目的:研究去甲异波尔定在大鼠肝微粒体中的葡萄糖醛酸化酶促反应动力学.方法:优化去甲异波尔定与大鼠肝微粒体的反应体系,采用超高效液相色谱-质谱联用技术定量检测孵育体系中去甲波尔定代谢产物去甲异波尔定-9-O-α-葡萄糖醛酸苷的浓度,并应用Linewearve-Burk作图分析数据,计算酶促动力学常数.结果:去甲异波尔定-9-O-α-葡萄糖醛酸苷的酶促反应动力学参数Km40.7μmol·L-1,Vmax=909.1 pmol·(min· mg pro)-1,肝清除率CLint(Vmax/Km)=22.3 μL· min-1·mg-1.结论:该方法简单、快速、可靠,适应于去甲异波尔定的葡萄糖醛酸化代谢研究;葡萄糖醛酸化是去甲异波尔定代谢的重要途径之一,提示葡萄糖醛酸转移酶的基因多态性及相关性的药物相互作用引起的去甲异波尔定活性和毒性作用的变化值得进一步关注.

  12. Classical mechanics without determinism

    OpenAIRE

    Nikolic, H.

    2005-01-01

    Classical statistical particle mechanics in the configuration space can be represented by a nonlinear Schrodinger equation. Even without assuming the existence of deterministic particle trajectories, the resulting quantum-like statistical interpretation is sufficient to predict all measurable results of classical mechanics. In the classical case, the wave function that satisfies a linear equation is positive, which is the main source of the fundamental difference between classical and quantum...

  13. Quantum computing classical physics.

    Science.gov (United States)

    Meyer, David A

    2002-03-15

    In the past decade, quantum algorithms have been found which outperform the best classical solutions known for certain classical problems as well as the best classical methods known for simulation of certain quantum systems. This suggests that they may also speed up the simulation of some classical systems. I describe one class of discrete quantum algorithms which do so--quantum lattice-gas automata--and show how to implement them efficiently on standard quantum computers.

  14. Complete kinetic and thermodynamic characterization of the unisite catalytic pathway of Escherichia coli F1-ATPase. Comparison with mitochondrial F1-ATPase and application to the study of mutant enzymes.

    Science.gov (United States)

    Al-Shawi, M K; Senior, A E

    1988-12-25

    A complete analysis is presented of the component rate constants of the "unisite" reaction pathway in normal Escherichia coli F1-ATPase. Gibbs free energy profiles of the unisite reaction pathway were constructed for both normal E. coli F1 and bovine-heart mitochondrial F1, and comparison indicated that E. coli F1 is an ancestral form of the mitochondrial enzyme. Similar kinetic and thermodynamic analyses of the unisite reaction pathway were done for mutant beta-Asn-242 and beta-Val-242 E. coli F1-ATPases. Both mutations affected unisite binding and hydrolysis of MgATP but had little effect on release of products or binding of MgADP. It was apparent that a primary effect of the mutations was on the interaction between the catalytic nucleotide-binding domain and the substrate MgATP. The catalytic transition state [F1-ATP]++ was the most destabilized step in the reaction sequence. Measurements of delta delta G[F1.ATP]++ and linear free energy plots for the catalytic step were consistent with the view that, in normal enzyme, residue beta-Asp-242 accepts an H-bond from the transition-state substrate in order to facilitate catalysis. Both mutations impaired positive catalytic cooperativity. This was caused by energetic destabilization of the catalytic transition state and was an indirect effect, not a direct effect on signal transmission per se between catalytic nucleotide-binding domains on beta-subunits. Therefore, impairment of unisite catalysis and of positive catalytic cooperativity appeared to be linked. This may provide a unifying explanation as to why a series of other, widely separated mis-sense mutations within the catalytic nucleotide-binding domain on F1-beta-subunit, which have been reported to affect unisite catalysis, also impair positive catalytic cooperativity. Linear free energy plots for the ATP-binding step of unisite catalysis demonstrated that beta-Asn-242 and beta-Val-242 mutant enzymes did not suffer any gross disruptive change in structure of

  15. ENZYME KINETICS OF APLYSIN METABOLISM IN RAT LIVER MICROSOMES%海兔素在大鼠肝微粒体中的酶促反应动力学研究

    Institute of Scientific and Technical Information of China (English)

    丰暖; 梁惠; 马爱国; 张辉珍

    2015-01-01

    Objective To establish a HPLC method for the determination of aplysin in rat liver microsome, and study the enzyme kinetics of aplysin metabolism in rat liver microsome, as well as the effect of CYP3A inhibitor ketoconazole. Methods A HPLC method was developed for the determination of aplysin. Aplysin was incubated in rat liver microsome for 30 min. The effects of different incubation times and liver microsomal protein concentrations on alysin metabolism were observed. Lineweave-Burk graphic method was used to calculate the enzyme kinetic parameters. Ketoconazole (0.5, 2, 5, 10, 20, 40 mg/L) was used to inhibit the metabolism of aplysin in vitro. Results The linear range of aplysia standard curve was 0.5-100 mg/L. The intra- and inter-day RSDs were less than 3.18%. The recovery rate was 96.19%-97.83%, and a good stability at room temperature for 24 h or at -200C for a week was confirmed. The best incubation conditions for aplysin metabolism in rat liver microsome were 30 min for incubation time and 1 mg/mL for protein concentration. The enzyme kinetic parameters were as follows: Km 19.72 mg/L, Vmax 0.35 mg/min/g protein, CLint 17.75 ml/(min·g) protein. Ketoconazole was inhibitory to the metabolism of aplysin in vitro. Conclusion The established HPLC method is fast, simple, specific and sensitive and applicable in the study of the enzyme kinetics of aplysin metabolism in rat liver microsome. CYP3A may be involved in the metabolism of aplysin.%目的建立测定大鼠肝微粒体孵育液中海兔素含量的高效液相色谱法(HPLC),研究海兔素在大鼠肝微粒体中代谢的酶动力学以及CYP3A特异性抑制剂酮康唑对其代谢反应的影响。方法考察不同孵育时间、肝微粒体蛋白浓度、海兔素浓度对代谢反应的影响,并采用双倒数作图法计算海兔素的酶促反应动力学参数;观察不同浓度酮康唑(0.5,2,5,10,20,40 mg/L)对海兔素体外代谢的影响。结果海兔素的

  16. Serodiagnosis of Lyme disease by kinetic enzyme-linked immunosorbent assay using recombinant VlsE1 or peptide antigens of Borrelia burgdorferi compared with 2-tiered testing using whole-cell lysates.

    Science.gov (United States)

    Bacon, Rendi Murphree; Biggerstaff, Brad J; Schriefer, Martin E; Gilmore, Robert D; Philipp, Mario T; Steere, Allen C; Wormser, Gary P; Marques, Adriana R; Johnson, Barbara J B

    2003-04-15

    In a study of US patients with Lyme disease, immunoglobulin (Ig) G and IgM antibody responses to recombinant Borrelia burgdorferi antigen VlsE1 (rVlsE1), IgG responses to a synthetic peptide homologous to a conserved internal sequence of VlsE (C6), and IgM responses to a synthetic peptide comprising the C-terminal 10 amino acid residues of a B. burgdorferi outer-surface protein C (pepC10) were evaluated by kinetic enzyme-linked immunoassay. At 99% specificity, the overall sensitivities for detecting IgG antibody to rVlsE1 or C6 in samples from patients with diverse manifestations of Lyme disease were equivalent to that of 2-tiered testing. When data were considered in parallel, 2 combinations (IgG responses to either rVlsE1 or C6 in parallel with IgM responses to pepC10) maintained high specificity (98%) and were significantly more sensitive than 2-tiered analysis in detecting antibodies to B. burgdorferi in patients with acute erythema migrans. In later stages of Lyme disease, the sensitivities of the in parallel tests and 2-tiered testing were high and statistically equivalent.

  17. 人肝微粒体中尼莫地平代谢的酶动力学及其抑制%Enzyme kinetics and inhibition of nimodipine metabolism in human liver microsomes

    Institute of Scientific and Technical Information of China (English)

    柳晓泉; 任亚丽; 钱之玉; 王广基

    2000-01-01

    AIM:To study the enzyme kinetics of nimodipine (NDP) metabolism and the effects of selective cytochrome P-450 (CYP-450) inhibitors on the metabolism of NDP in human liver microsomes in vitro. METHODS: Microsomes from six individual human liver specimens were used to perform enzyme kinetic studies and the kinetic parameters were estimated by Eadie-Hofstee equation. Various selective CYP-450 inhibitors were used to investigate their effects on the metabolism of NDP and the principal CYP-450 isoform involved in dehydrogenation of dihydropyridine ring of NDP in human liver microsomes. RESULTS: There was an important intersubject variability in NDP metabolism in human liver microsomes. For NDP dehydrogeuase activity, the Km value was (36 + 11 ) μmol and the Vm value was ( 17 + 7) μmol · g- 1· min- 1. The dehydrogenation of dihydropyridine ring of NDP was competitively inhibited by ketoconazole (Ket) and troleandomycin (TAO), and the Ki values for Ket and TAO were 0.59 and 122.2 μmol, respectively.Phenacetin ( Pnt ), quinidine ( Qui ), diethyldithiocarbamate (DDC), sulfaphenazole (Sul), and tranylcypromine (Tta) had a little or no inhibitory effects on the dehydrogenation of NDP. CONCLUSION: The intersubject variability of NDP pharmacokinetics was attributed to the metabolic polymorphism of NDP in liver.Cytochrome P-4503A (CYP3A) is involved in the dehydrogenation of dihydropyridine ring of NDP.%目的:在体外研究人肝微粒体中尼莫地平代谢的酶动力学及选择性的细胞色素P-450(CYP450)酶抑制剂对尼莫地平代谢的影响.方法:采用人肝微粒体研究尼莫地平脱氢酶的代谢动力学,运用Eadie-Hof-stee方程估算其动力学参数.在体外运用CYP450酶的选择性抑制剂探讨其对尼莫地平代谢的影响及人肝微粒体中参与尼莫地平二氢吡啶环脱氢代谢的CYP450酶.结果:尼莫地平在人肝微粒体中的代谢存在较大的个体差异,尼莫地平脱氢酶的Km为(36±11)

  18. Entanglement in Classical Optics

    CERN Document Server

    Ghose, Partha

    2013-01-01

    The emerging field of entanglement or nonseparability in classical optics is reviewed, and its similarities with and differences from quantum entanglement clearly pointed out through a recapitulation of Hilbert spaces in general, the special restrictions on Hilbert spaces imposed in quantum mechanics and the role of Hilbert spaces in classical polarization optics. The production of Bell-like states in classical polarization optics is discussed, and new theorems are proved to discriminate between separable and nonseparable states in classical wave optics where no discreteness is involved. The influence of the Pancharatnam phase on a classical Bell-like state is deived. Finally, to what extent classical polarization optics can be used to simulate quantum information processing tasks is also discussed. This should be of great practical importance because coherence and entanglement are robust in classical optics but not in quantum systems.

  19. Measuring the Enzyme Activity of Arabidopsis Deubiquitylating Enzymes.

    Science.gov (United States)

    Kalinowska, Kamila; Nagel, Marie-Kristin; Isono, Erika

    2016-01-01

    Deubiquitylating enzymes, or DUBs, are important regulators of ubiquitin homeostasis and substrate stability, though the molecular mechanisms of most of the DUBs in plants are not yet understood. As different ubiquitin chain types are implicated in different biological pathways, it is important to analyze the enzyme characteristic for studying a DUB. Quantitative analysis of DUB activity is also important to determine enzyme kinetics and the influence of DUB binding proteins on the enzyme activity. Here, we show methods to analyze DUB activity using immunodetection, Coomassie Brilliant Blue staining, and fluorescence measurement that can be useful for understanding the basic characteristic of DUBs.

  20. Electrogravimetric real-time and in situ michaelis-menten enzymatic kinetics: progress curve of acetylcholinesterase hydrolysis.

    Science.gov (United States)

    Bueno, Paulo R; Watanabe, Ailton M; Faria, Ronaldo C; Santos, Márcio L; Riccardi, Carla S

    2010-12-16

    A piezoelectric detection of enzyme-modified surface was performed under Michaelis-Menten presumptions of steady-state condition. The approach herein presented showed promise in the study of enzymatic kinetics by measuring the frequency changes associated with mass changes at the piezoelectric crystal surface. Likewise, real-time frequency shifts, that is, dΔf/dt, indicated the rate of products formation from enzymatic reaction. In this paper, acetylcholinesterase was used as the enzymatic model and acetylcholine as substrate. The enzymatic rate has its maximum value for a short time during the kinetic reaction, for instance, during the first ten minutes of the reaction time scale. The values found for the kinetic constant rate and Michaelis-Menten constant were (1.4 ± 0.8) 10(5) s(-1) and (5.2 ± 3) 10(-4) M, respectively, in agreement with the values found in classical Michaelis-Menten kinetic experiments.

  1. Adsorption studies of molasse's wastewaters on activated carbon: modelling with a new fractal kinetic equation and evaluation of kinetic models.

    Science.gov (United States)

    Figaro, S; Avril, J P; Brouers, F; Ouensanga, A; Gaspard, S

    2009-01-30

    Adsorption kinetic of molasses wastewaters after anaerobic digestion (MSWD) and melanoidin respectively on activated carbon was studied at different pH. The kinetic parameters could be determined using classical kinetic equations and a recently published fractal kinetic equation. A linear form of this equation can also be used to fit adsorption data. Even with lower correlation coefficients the fractal kinetic equation gives lower normalized standard deviation values than the pseudo-second order model generally used to fit adsorption kinetic data, indicating that the fractal kinetic model is much more accurate for describing the kinetic adsorption data than the pseudo-second order kinetic model.

  2. Classical, Semi-classical and Quantum Noise

    CERN Document Server

    Poor, H; Scully, Marlan

    2012-01-01

    David Middleton was a towering figure of 20th Century engineering and science and one of the founders of statistical communication theory. During the second World War, the young David Middleton, working with Van Fleck, devised the notion of the matched filter, which is the most basic method used for detecting signals in noise. Over the intervening six decades, the contributions of Middleton have become classics. This collection of essays by leading scientists, engineers and colleagues of David are in his honor and reflect the wide  influence that he has had on many fields. Also included is the introduction by Middleton to his forthcoming book, which gives a wonderful view of the field of communication, its history and his own views on the field that he developed over the past 60 years. Focusing on classical noise modeling and applications, Classical, Semi-Classical and Quantum Noise includes coverage of statistical communication theory, non-stationary noise, molecular footprints, noise suppression, Quantum e...

  3. Thermodynamics and kinetics of an enzyme-catalyzed reaction determined by isothermal titration calorimetry%等温滴定量热法测定酶催化反应的热动力学参数

    Institute of Scientific and Technical Information of China (English)

    彭尚; 孙丽霞; 熊珍爱; 周利琴; 兰雄雕; 孙建华; 童张法; 廖丹葵

    2016-01-01

    采用等温滴定量热法(ITC)测定猪肺血管紧张素转化酶(angiotensin converting enzyme,ACE)催化水解其体外模拟底物马尿酰-组氨酰-亮氨酸(Hip-His-Leu,HHL)反应的热动力学参数,考察了温度对动力学参数的影响。结果表明,该反应的摩尔水解焓ΔHhydr为正值,是吸热反应,且随温度升高ΔHhydr增大,等压比热容 cp为0.2126kJ/(mol·K);ACE 催化 HHL 的水解反应符合 Michaelis-Menten 机理,在实验温度范围内(298.15~313.15K),米氏常数Km随温度升高而减小,催化常数kcat随温度的升高先增大后减少,在308.15K时达到最大值2.534s−1。将该法与传统的初始速率法进行比较,传统法存在的局限性使测得的 Km相对偏大。同时使用 ITC结合动力学分析测得ACE抑制剂药物依那普利拉为竞争性抑制剂,抑制常数KI为12.1 nmol/L,与文献比较证明该法可用于抑制剂类型的判断,是一种开发 ACE 抑制剂的新方法。应用该方法确定活性多肽Arg-Tyr-Leu-Gly-Tyr(RY-5)为非竞争性抑制剂,抑制常数KI为1.0μmol/L。%Thermodynamic and kinetic parameters of angiotensin converting enzyme(ACE)catalyzed hydrolysis of simulating substrate Hippuryl-Histidyl-Leucine(HHL)in vitro were determined by isothermal titration calorimetry(ITC). The effect of temperature on kinetic parameters was investigated; the results showed that the ACE-catalyzed reaction was endothermic with a small constant pressure specific heat capacity [cp=0.2126kJ/(mol·K)]. The value of molar hydrolysis enthalpyΔHhydrwas positive and increased as temperature rose. The reaction mechanism was in accordance with the Michaelis-Menten model in the temperature range(298.15—313.15K);the effect of temperature on the Michaelis constant(Km)was negative,while catalytic constant(kcat)first increased then decreased with the increase of temperature,reaching the maximum value of 2.534s−1 at 308.15K. Initial rate

  4. Introducing Michaelis-Menten Kinetics through Simulation

    Science.gov (United States)

    Halkides, Christopher J.; Herman, Russell

    2007-01-01

    We describe a computer tutorial that introduces the concept of the steady state in enzyme kinetics. The tutorial allows students to produce graphs of the concentrations of free enzyme, enzyme-substrate complex, and product versus time in order to learn about the approach to steady state. By using a range of substrate concentrations and rate…

  5. Controlled enzyme catalyzed heteropolysaccharide degradation

    DEFF Research Database (Denmark)

    Rasmussen, Louise Enggaard

    The work presented in this PhD thesis has provided a better understanding of the enzyme kinetics and quantitative phenomena of the hydrolysis of xylan substrates by selected pure enzyme preparations. Furthermore, the options for producing specific substituted xylooligosaccharides from selected...... substrates by specific xylanase treatment have been examined. The kinetics of the enzymatic degradation of water-extractable wheat arabinoxylan (WE-AX) during designed treatments with selected monocomponent enzymes was investigated by monitoring the release of xylose and arabinose. The results of different...... between -xylosidase and the α-L-arabinofuranosidases on the xylose release were low as compared to the effect of xylanase addition with β-xylosidase, which increased the xylose release by ~25 times in 30 minutes. At equimolar addition levels of the four enzymes, the xylanase activity was thus rate...

  6. Lectures on Classical Integrability

    CERN Document Server

    Torrielli, Alessandro

    2016-01-01

    We review some essential aspects of classically integrable systems. The detailed outline of the lectures consists of: 1. Introduction and motivation, with historical remarks; 2. Liouville theorem and action-angle variables, with examples (harmonic oscillator, Kepler problem); 3. Algebraic tools: Lax pairs, monodromy and transfer matrices, classical r-matrices and exchange relations, non-ultralocal Poisson brackets, with examples (non-linear Schroedinger model, principal chiral field); 4. Features of classical r-matrices: Belavin-Drinfeld theorems, analyticity properties, and lift of the classical structures to quantum groups; 5. Classical inverse scattering method to solve integrable differential equations: soliton solutions, spectral properties and the Gel'fand-Levitan-Marchenko equation, with examples (KdV equation, Sine-Gordon model). Prepared for the Durham Young Researchers Integrability School, organised by the GATIS network. This is part of a collection of lecture notes.

  7. 苯甘氨酸氨基转移酶基因hpgt的原核优化表达与酶动力学特性研究%Optimized Prokaryotic Expression of the Recombinant hpgt Gene and Enzyme Kinetic Characteristics

    Institute of Scientific and Technical Information of China (English)

    王宗瑞; 赵广荣

    2012-01-01

    4-Hydroxyphenylglycine aminotransferase which can synthesize D-phenylglycine transaminase is produced by Pseudomonas. The hpgt gene was synthesized through the codon optimization technology. The recombinant prokaryotic plasmid pCDF-hpgt was obtained. The plasmid was transformed into the competent cell E. coli BL21 (DE3). The recombinant His-HpgT protein was obtained after the optimized expression and purified by nickel chelate affinity chromatography method. The enzyme activity of the forward and reverse reactions was measured and the activity of the forward reaction reached 749mU/mg which was lower than the reverse, 2 257mU/mg. Also the optimized temperature and pH were measured, with the result of 35癚.anA 8.0. Other kinetic parameters and the mechanism analysis of enzyme characteristics were explained. The enzyme affinity to phenylglycine was higher than the glutamate which obtained by the Michaelis-Menten equation; and the reaction was inhibited by the lower concentration of phenylglyoxylic acid.%苯甘氨酸氨基转移酶(4-Hydroxyphenylglycine aminotransferase)是假单胞菌所产生的一种能够合成D-苯甘氨酸的重要转氨酶.利用密码子优化技术,合成苯甘氨酸转移酶基因.构建原核重组质粒pCDF-hpgt,转入感受态细胞E.coli BL21 (DE3),优化表达His-HpgT蛋白.利用Ni-NTA柱纯化技术获得高纯度的His-HpgT融合蛋白.分别测定融合蛋白在正反向反应中的酶活力单位及最佳的反应温度、pH值及其他动力学参数,并对该酶特性作相关的机理分析.测定结果表明,正向反应和反向反应的酶比活力分别为749mU/mg、2 257mU/mg,此酶分解苯甘氨酸的能力要强于合成苯甘氨酸;正向反应的最适温度与pH分别是35℃和8.0;由米氏方程得出该酶对苯甘氨酸的亲和力远大于谷氨酸;较低浓度的苯乙醛酸即可抑制反应的进行.

  8. Kinetic theory of nonideal gases and nonideal plasmas

    CERN Document Server

    Klimontovich, Yu L

    2013-01-01

    Kinetic Theory of Nonideal Gases and Nonideal Plasmas presents the fundamental aspects of the kinetic theory of gases and plasmas. The book consists of three parts, which attempts to present some of the ideas, methods and applications in the study of the kinetic processes in nonideal gases and plasmas. The first part focuses on the classical kinetic theory of nonideal gases. The second part discusses the classical kinetic theory of fully ionized plasmas. The last part is devoted to the quantum kinetic theory of nonideal gases and plasmas. A concluding chapter is included, which presents a shor

  9. Enzyme Molecules in Solitary Confinement

    Directory of Open Access Journals (Sweden)

    Raphaela B. Liebherr

    2014-09-01

    Full Text Available Large arrays of homogeneous microwells each defining a femtoliter volume are a versatile platform for monitoring the substrate turnover of many individual enzyme molecules in parallel. The high degree of parallelization enables the analysis of a statistically representative enzyme population. Enclosing individual enzyme molecules in microwells does not require any surface immobilization step and enables the kinetic investigation of enzymes free in solution. This review describes various microwell array formats and explores their applications for the detection and investigation of single enzyme molecules. The development of new fabrication techniques and sensitive detection methods drives the field of single molecule enzymology. Here, we introduce recent progress in single enzyme molecule analysis in microwell arrays and discuss the challenges and opportunities.

  10. Enzyme molecules in solitary confinement.

    Science.gov (United States)

    Liebherr, Raphaela B; Gorris, Hans H

    2014-09-12

    Large arrays of homogeneous microwells each defining a femtoliter volume are a versatile platform for monitoring the substrate turnover of many individual enzyme molecules in parallel. The high degree of parallelization enables the analysis of a statistically representative enzyme population. Enclosing individual enzyme molecules in microwells does not require any surface immobilization step and enables the kinetic investigation of enzymes free in solution. This review describes various microwell array formats and explores their applications for the detection and investigation of single enzyme molecules. The development of new fabrication techniques and sensitive detection methods drives the field of single molecule enzymology. Here, we introduce recent progress in single enzyme molecule analysis in microwell arrays and discuss the challenges and opportunities.

  11. Kinetic Typography

    DEFF Research Database (Denmark)

    van Leeuwen, Theo; Djonov, Emilia

    2014-01-01

    After discussing broad cultural drivers behind the development of kinetic typography, the chapter outlines an approach to analysing kinetic typography which is based on Halliday's theory of transitivity, as applied by Kress and Van Leeuwen to visual images.......After discussing broad cultural drivers behind the development of kinetic typography, the chapter outlines an approach to analysing kinetic typography which is based on Halliday's theory of transitivity, as applied by Kress and Van Leeuwen to visual images....

  12. 21 CFR 862.2500 - Enzyme analyzer for clinical use.

    Science.gov (United States)

    2010-04-01

    ... (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Laboratory... use is a device intended to measure enzymes in plasma or serum by nonkinetic or kinetic measurement...

  13. Enzymes: principles and biotechnological applications.

    Science.gov (United States)

    Robinson, Peter K

    2015-01-01

    Enzymes are biological catalysts (also known as biocatalysts) that speed up biochemical reactions in living organisms, and which can be extracted from cells and then used to catalyse a wide range of commercially important processes. This chapter covers the basic principles of enzymology, such as classification, structure, kinetics and inhibition, and also provides an overview of industrial applications. In addition, techniques for the purification of enzymes are discussed.

  14. Enzymatic Kinetic Isotope Effects from Path-Integral Free Energy Perturbation Theory.

    Science.gov (United States)

    Gao, J

    2016-01-01

    Path-integral free energy perturbation (PI-FEP) theory is presented to directly determine the ratio of quantum mechanical partition functions of different isotopologs in a single simulation. Furthermore, a double averaging strategy is used to carry out the practical simulation, separating the quantum mechanical path integral exactly into two separate calculations, one corresponding to a classical molecular dynamics simulation of the centroid coordinates, and another involving free-particle path-integral sampling over the classical, centroid positions. An integrated centroid path-integral free energy perturbation and umbrella sampling (PI-FEP/UM, or simply, PI-FEP) method along with bisection sampling was summarized, which provides an accurate and fast convergent method for computing kinetic isotope effects for chemical reactions in solution and in enzymes. The PI-FEP method is illustrated by a number of applications, to highlight the computational precision and accuracy, the rule of geometrical mean in kinetic isotope effects, enhanced nuclear quantum effects in enzyme catalysis, and protein dynamics on temperature dependence of kinetic isotope effects.

  15. 补骨脂素和异补骨脂素在大鼠和人肝微粒体的酶促动力学%Enzyme kinetics of psoralen and isopsoralen in rat and human liver microsomes

    Institute of Scientific and Technical Information of China (English)

    阳海鹰; 钟玉环; 陈琳; 李桦; 庄笑梅

    2015-01-01

    目的:研究中药有效成分补骨脂素和异补骨脂素的细胞色素P450(CYP)酶促动力学特征,比较其结构和种属差异,为体内药代动力学特征的预测提供科学依据。方法建立补骨脂素和异补骨脂素的液相色谱串联质谱(LC-MS/MS)检测方法,在优化人和大鼠肝微粒体孵育体系和评价体外代谢稳定性的基础上,进行了代谢稳定性和酶促动力学研究,应用非线性回归法计算最大反应速率(Vmax)和米氏常数(Km)。结果应用建立的LC-MS/MS检测方法,补骨脂素、异补骨脂素在肝微粒体孵育液中定量范围为0.1~50.0μmol·L-1,线性关系良好,精密度和准确度等满足检测要求。体外代谢稳定性研究显示,当底物浓度为1μmol·L-1,蛋白浓度为0.5 g·L-1,孵育40 min内,补骨脂素和异补骨脂素在大鼠和人肝微粒体呈线性消除,体外半衰期分别为74.5,95.0,74.5和173.3 min。补骨脂素在大鼠和人肝微粒体中的Vmax分别为:(1.140±0.080)μmol·min-1·g-1蛋白,(0.620±0.060)μmol·min-1·g-1蛋白;Km分别为(12.9±0.3)μmol·L-1和(7.4±1.3)μmol·L-1。异补骨脂素在大鼠和人肝微粒体中的Vmax分别为(0.251±0.012)μmol·min-1·g-1蛋白和(0.103±0.014)μmol·min-1·g-1蛋白;Km分别为:(3.0±0.4)μmol·L-1和(3.4±0.7)μmol·L-1。结论补骨脂素和异补骨脂素的CYP酶促动力学特征及代谢稳定性具有一定的种属和结构差异,在大鼠两者基于CYP酶的代谢清除过程可能相似。而在人体,异补骨脂素的CYP代谢清除可能慢于补骨脂素,导致药代动力学特征的差异。%OBJECTIVE To investigate and compare the enzyme kinetic characters of psoralen (PRN)and isopsoralen(IPRN)in rat and human liver microsomes. METHODS PRN and IPRN in liver microsomes incubates were determined using LC-MS/MS. The enzyme kinetic and metabolic stability of PRN and IPRN were

  16. Characterization of CIM monoliths as enzyme reactors.

    Science.gov (United States)

    Vodopivec, Martina; Podgornik, Ales; Berovic, Marin; Strancar, Ales

    2003-09-25

    The immobilization of the enzymes citrate lyase, malate dehydrogenase, isocitrate dehydrogenase and lactate dehydrogenase to CIM monolithic supports was performed. The long-term stability, reproducibility, and linear response range of the immobilized enzyme reactors were investigated along with the determination of the kinetic behavior of the enzymes immobilized on the CIM monoliths. The Michaelis-Menten constant K(m) and the turnover number k(3) of the immobilized enzymes were found to be flow-unaffected. Furthermore, the K(m) values of the soluble and immobilized enzyme were found to be comparable. Both facts indicate the absence of a diffusional limitation in immobilized CIM enzyme reactors.

  17. Fermions from classical statistics

    OpenAIRE

    2010-01-01

    We describe fermions in terms of a classical statistical ensemble. The states $\\tau$ of this ensemble are characterized by a sequence of values one or zero or a corresponding set of two-level observables. Every classical probability distribution can be associated to a quantum state for fermions. If the time evolution of the classical probabilities $p_\\tau$ amounts to a rotation of the wave function $q_\\tau(t)=\\pm \\sqrt{p_\\tau(t)}$, we infer the unitary time evolution of a quantum system of fe...

  18. Davidson and classical pragmatism

    Directory of Open Access Journals (Sweden)

    Paula Rossi

    2007-06-01

    Full Text Available In this paper I wish to trace some connections between Donald Davidson's work (1917-2003 and two major representatives of the classical pragmatist movement: Charles S. Peirce (1839-1914 and William James (1842-1910. I will start with a basic characterization of classical pragmatism; then, I shall examine certain conceptions in Peirce's and James' pragmatism, in order to establish affinities with Davidson´s thought. Finally, and bearing in mind the previous con-nections, I will reflect briefly on the relevance –often unrecognized- of classical pragmatist ideas in the context of contemporary philosophi-cal discussions.

  19. Kinetic analysis of Enterococcus faecium L,D-transpeptidase inactivation by carbapenems.

    Science.gov (United States)

    Dubée, Vincent; Arthur, Michel; Fief, Hélène; Triboulet, Sébastien; Mainardi, Jean-Luc; Gutmann, Laurent; Sollogoub, Matthieu; Rice, Louis B; Ethève-Quelquejeu, Mélanie; Hugonnet, Jean-Emmanuel

    2012-06-01

    Bypass of classical penicillin-binding proteins by the L,D-transpeptidase of Enterococcus faecium (Ldt(fm)) leads to high-level ampicillin resistance in E. faecium mutants, whereas carbapenems remain the lone highly active β-lactams. Kinetics of Ldt(fm) inactivation was determined for four commercial carbapenems and a derivative obtained by introducing a minimal ethyl group at position 2. We show that the bulky side chains of commercial carbapenems have both positive and negative effects in preventing hydrolysis of the acyl enzyme and impairing drug binding.

  20. Comparison on Metabolic Enzyme Kinetics of Puerarin and Its Crude Extract in Rat Liver Microsomes%葛根素及葛根提取物在大鼠肝微粒体内代谢动力学研究

    Institute of Scientific and Technical Information of China (English)

    刘莹; 王国丽

    2011-01-01

    To compare the differences of enzyme kinetics of puerarin and its crude extract metabolism via rat liver microsomes, a HPLC method was established for quantitative analysis of puerarin in liver microsomes incubation system with p-hydroxy benzoic acid as internal standard. An optimum incubation system was found and the parameters of kinetics were analyzed and calculated with the software of CraphPad Prism 5.0. In the metabolic system,the optimum incubated time was 20 min.and the optimum protein concentration of rat liver microsomes was 1.0 mg/mL. The Km, Vmax and Clint ( Vmax/Km) of monomeric puerarin were (0. 93 ±0. 17) μmol/L,(93.59 ±2. 82) nmol/min'mg and (97.49 ±1.07) mL/min'mg, respectively. Those of puerarin in crude extract were (1.02 ±0.09) u,mol/L, (22.48 ± 1. 53) nmol/min-mg and (22. 04 ± 1. 72) mL/min'mg, respectively. The Km values of two kinds of puerarin were similar,but their Vmax and Clint had significant differences. The Vmax values varied by 4.16 folds and the Clint values differed by 4.42 fold. The results showed that the metabolism of puerarin was faster than that of puerarin in crude extract via rat liver microsomes. Other components in Chinese herbs may influence the metabolism of the active ones.%为了研究葛根素单体和葛根提取物中的葛根素在大鼠肝微粒体中的代谢是否存在差异,建立微粒体孵育体系.采用高效液相色谱法测定葛根素在肝微粒体代谢系统中的含量,并应用GraphPad Prism 5.0软件分析数据,计算酶促反应动力学参数.在体外代谢系统中,葛根素单体的酶促反应动力学参数Km为(0.93±0.17)μmol/L,Vmax为(93.59±2.82) nmol/min·mg,CLint(Vmax/Km)为(100.6±1.07) mL/min·mg;葛根提取物中葛根素的相应的动力学参数分别为(1.02±0.09)μmol/L,(22.48±1.53) nmol/min· mg和(22.04±1.72) mL/min·mg.葛根素单体和葛根提取物中的葛根素在大鼠肝微粒体的代谢存在明显的差异,葛根素单体的代谢速度快于

  1. Advanced classical field theory

    CERN Document Server

    Giachetta, Giovanni; Sardanashvily, Gennadi

    2009-01-01

    Contemporary quantum field theory is mainly developed as quantization of classical fields. Therefore, classical field theory and its BRST extension is the necessary step towards quantum field theory. This book aims to provide a complete mathematical foundation of Lagrangian classical field theory and its BRST extension for the purpose of quantization. Based on the standard geometric formulation of theory of nonlinear differential operators, Lagrangian field theory is treated in a very general setting. Reducible degenerate Lagrangian theories of even and odd fields on an arbitrary smooth manifold are considered. The second Noether theorems generalized to these theories and formulated in the homology terms provide the strict mathematical formulation of BRST extended classical field theory

  2. On Noncommutative Classical Mechanics

    CERN Document Server

    Djemai, A E F

    2003-01-01

    In this work, I investigate the noncommutative Poisson algebra of classical observables corresponding to a proposed general Noncommutative Quantum Mechanics, \\cite{1}. I treat some classical systems with various potentials and some Physical interpretations are given concerning the presence of noncommutativity at large scales (Celeste Mechanics) directly tied to the one present at small scales (Quantum Mechanics) and its possible relation with UV/IR mixing.

  3. Between classical and quantum

    OpenAIRE

    2007-01-01

    The relationship between classical and quantum theory is of central importance to the philosophy of physics, and any interpretation of quantum mechanics has to clarify it. Our discussion of this relationship is partly historical and conceptual, but mostly technical and mathematically rigorous, including over 500 references. On the assumption that quantum mechanics is universal and complete, we discuss three ways in which classical physics has so far been believed to emerge from quantum physic...

  4. Quantum and Classic Brackets

    OpenAIRE

    Kisil, Vladimir V.

    2000-01-01

    We describe an $p$-mechanical (see funct-an/9405002 and quant-ph/9610016) brackets which generate quantum (commutator) and classic (Poisson) brackets in corresponding representations of the Heisenberg group. We \\emph{do not} use any kind of semiclassic approximation or limiting procedures for $\\hbar \\to 0$. Harmonic oscillator considered within the approach. Keywords: Classic and quantum mechanics, Hamilton and Heisenberg equations, Poisson brackets, commutator, Heisenberg group.

  5. In vivo doses of butadiene epoxides as estimated from in vitro enzyme kinetics by using cob(I)alamin and measured hemoglobin adducts: An inter-species extrapolation approach

    Energy Technology Data Exchange (ETDEWEB)

    Motwani, Hitesh V., E-mail: hitesh.motwani@mmk.su.se; Törnqvist, Margareta

    2014-12-15

    1,3-Butadiene (BD) is a rodent and human carcinogen. In the cancer tests, mice have been much more susceptible than rats with regard to BD-induced carcinogenicity. The species-differences are dependent on metabolic formation/disappearance of the genotoxic BD epoxy-metabolites that lead to variations in the respective in vivo doses, i.e. “area under the concentration-time curve” (AUC). Differences in AUC of the most gentoxic BD epoxy-metabolite, diepoxybutane (DEB), are considered important with regard to cancer susceptibility. The present work describes: the application of cob(I)alamin for accurate measurements of in vitro enzyme kinetic parameters associated with BD epoxy-metabolites in human, mouse and rat; the use of published data on hemoglobin (Hb) adduct levels of BD epoxides from BD exposure studies on the three species to calculate the corresponding AUCs in blood; and a parallelogram approach for extrapolation of AUC of DEB based on the in vitro metabolism studies and adduct data from in vivo measurements. The predicted value of AUC of DEB for humans from the parallelogram approach was 0.078 nM · h for 1 ppm · h of BD exposure compared to 0.023 nM · h/ppm · h as calculated from Hb adduct levels observed in occupational exposure. The corresponding values in nM · h/ppm · h were for mice 41 vs. 38 and for rats 1.26 vs. 1.37 from the parallelogram approach vs. experimental exposures, respectively, showing a good agreement. This quantitative inter-species extrapolation approach will be further explored for the clarification of metabolic rates/pharmacokinetics and the AUC of other genotoxic electrophilic compounds/metabolites, and has a potential to reduce and refine animal experiments. - Highlights: • In vitro metabolism to in vivo dose extrapolation of butadiene metabolites was proposed. • A parallelogram approach was introduced to estimate dose (AUC) in humans and rodents. • AUC of diepoxybutane predicted in humans was 0.078 nM h/ppm h

  6. Michaelis-Menten kinetics under spatially constrained conditions: application to mibefradil pharmacokinetics.

    Science.gov (United States)

    Kosmidis, Kosmas; Karalis, Vangelis; Argyrakis, Panos; Macheras, Panos

    2004-09-01

    Two different approaches were used to study the kinetics of the enzymatic reaction under heterogeneous conditions to interpret the unusual nonlinear pharmacokinetics of mibefradil. Firstly, a detailed model based on the kinetic differential equations is proposed to study the enzymatic reaction under spatial constraints and in vivo conditions. Secondly, Monte Carlo simulations of the enzyme reaction in a two-dimensional square lattice, placing special emphasis on the input and output of the substrate were applied to mimic in vivo conditions. Both the mathematical model and the Monte Carlo simulations for the enzymatic reaction reproduced the classical Michaelis-Menten (MM) kinetics in homogeneous media and unusual kinetics in fractal media. Based on these findings, a time-dependent version of the classic MM equation was developed for the rate of change of the substrate concentration in disordered media and was successfully used to describe the experimental plasma concentration-time data of mibefradil and derive estimates for the model parameters. The unusual nonlinear pharmacokinetics of mibefradil originates from the heterogeneous conditions in the reaction space of the enzymatic reaction. The modified MM equation can describe the pharmacokinetics of mibefradil as it is able to capture the heterogeneity of the enzymatic reaction in disordered media.

  7. Moonlighting enzymes in parasitic protozoa.

    Science.gov (United States)

    Collingridge, Peter W; Brown, Robert W B; Ginger, Michael L

    2010-08-01

    Enzymes moonlight in a non-enzymatic capacity in a diverse variety of cellular processes. The discovery of these non-enzymatic functions is generally unexpected, and moonlighting enzymes are known in both prokaryotes and eukaryotes. Importantly, this unexpected multi-functionality indicates that caution might be needed on some occasions in interpreting phenotypes that result from the deletion or gene-silencing of some enzymes, including some of the best known enzymes from classic intermediary metabolism. Here, we provide an overview of enzyme moonlighting in parasitic protists. Unequivocal and putative examples of moonlighting are discussed, together with the possibility that the unusual biological characteristics of some parasites either limit opportunities for moonlighting to arise or perhaps contribute to the evolution of novel proteins with clear metabolic ancestry.

  8. Kinetic theory and transport phenomena

    CERN Document Server

    Soto, Rodrigo

    2016-01-01

    This textbook presents kinetic theory, which is a systematic approach to describing nonequilibrium systems. The text is balanced between the fundamental concepts of kinetic theory (irreversibility, transport processes, separation of time scales, conservations, coarse graining, distribution functions, etc.) and the results and predictions of the theory, where the relevant properties of different systems are computed. The book is organised in thematic chapters where different paradigmatic systems are studied. The specific features of these systems are described, building and analysing the appropriate kinetic equations. Specifically, the book considers the classical transport of charges, the dynamics of classical gases, Brownian motion, plasmas, and self-gravitating systems, quantum gases, the electronic transport in solids and, finally, semiconductors. Besides these systems that are studied in detail, concepts are applied to some modern examples including the quark–gluon plasma, the motion of bacterial suspen...

  9. Enzyme technology: Key to selective biorefining

    DEFF Research Database (Denmark)

    Meyer, Anne S.

    2014-01-01

    to the reaction is a unique trait of enzyme catalysis. Since enzyme selectivity means that a specific reaction is catalysed between particular species to produce definite products, enzymes are particularly fit for converting specific compounds in mixed biomass streams. Since enzymes are protein molecules...... their rational use in biorefinery processes requires an understanding of the basic features of enzymes and reaction traits with respect to specificity, kinetics, reaction optima, stability and structure-function relations – we are now at a stage where it is possible to use nature’s enzyme structures as starting...... point and then improve the functional traits by targeted mutation of the protein. The talk will display some of our recent hypotheses related to enzyme action, recently obtained results within knowledge-based enzyme improvements as well as cast light on research methods used in optimizing enzyme...

  10. Enzymes in CO2 Capture

    DEFF Research Database (Denmark)

    Fosbøl, Philip Loldrup; Gladis, Arne; Thomsen, Kaj

    of carbon capture is the application of enzymes for acceleration of typically slow ternary amines or inorganic carbonates. There is a hidden potential to revive currently infeasible amines which have an interesting low energy consumption for regeneration but too slow kinetics for viable CO2 capture. The aim......The enzyme Carbonic Anhydrase (CA) can accelerate the absorption rate of CO2 into aqueous solutions by several-fold. It exist in almost all living organisms and catalyses different important processes like CO2 transport, respiration and the acid-base balances. A new technology in the field...... of this work is to discuss the measurements of kinetic properties for CA promoted CO2 capture solvent systems. The development of a rate-based model for enzymes will be discussed showing the principles of implementation and the results on using a well-known ternary amine for CO2 capture. Conclusions...

  11. Halophilic adaptation of enzymes.

    Science.gov (United States)

    Madern, D; Ebel, C; Zaccai, G

    2000-04-01

    It is now clear that the understanding of halophilic adaptation at a molecular level requires a strategy of complementary experiments, combining molecular biology, biochemistry, and cellular approaches with physical chemistry and thermodynamics. In this review, after a discussion of the definition and composition of halophilic enzymes, the effects of salt on their activity, solubility, and stability are reviewed. We then describe how thermodynamic observations, such as parameters pertaining to solvent-protein interactions or enzyme-unfolding kinetics, depend strongly on solvent composition and reveal the important role played by water and ion binding to halophilic proteins. The three high-resolution crystal structures now available for halophilic proteins are analyzed in terms of haloadaptation, and finally cellular response to salt stress is discussed briefly.

  12. Illustrating Enzyme Inhibition Using Gibbs Energy Profiles

    Science.gov (United States)

    Bearne, Stephen L.

    2012-01-01

    Gibbs energy profiles have great utility as teaching and learning tools because they present students with a visual representation of the energy changes that occur during enzyme catalysis. Unfortunately, most textbooks divorce discussions of traditional kinetic topics, such as enzyme inhibition, from discussions of these same topics in terms of…

  13. Enzyme Catalysis and the Gibbs Energy

    Science.gov (United States)

    Ault, Addison

    2009-01-01

    Gibbs-energy profiles are often introduced during the first semester of organic chemistry, but are less often presented in connection with enzyme-catalyzed reactions. In this article I show how the Gibbs-energy profile corresponds to the characteristic kinetics of a simple enzyme-catalyzed reaction. (Contains 1 figure and 1 note.)

  14. Discrete Classical Electromagnetic Fields

    CERN Document Server

    De Souza, M M

    1997-01-01

    The classical electromagnetic field of a spinless point electron is described in a formalism with extended causality by discrete finite transverse point-vector fields with discrete and localized point interactions. These fields are taken as a classical representation of photons, ``classical photons". They are all transversal photons; there are no scalar nor longitudinal photons as these are definitely eliminated by the gauge condition. The angular distribution of emitted photons coincides with the directions of maximum emission in the standard formalism. The Maxwell formalism and its standard field are retrieved by the replacement of these discrete fields by their space-time averages, and in this process scalar and longitudinal photons are necessarily created and added. Divergences and singularities are by-products of this averaging process. This formalism enlighten the meaning and the origin of the non-physical photons, the ones that violate the Lorentz condition in manifestly covariant quantization methods.

  15. Between classical and quantum

    CERN Document Server

    Landsman, N P

    2005-01-01

    The relationship between classical and quantum theory is of central importance to the philosophy of physics, and any interpretation of quantum mechanics has to clarify it. Our discussion of this relationship is partly historical and conceptual, but mostly technical and mathematically rigorous, including over 500 references. On the assumption that quantum mechanics is universal and complete, we discuss three ways in which classical physics has so far been believed to emerge from quantum physics, namely in the limit h -> 0 of small Planck's constant (in a finite system), in the limit of a large system, and through decoherence and consistent histores. The first limit is closely related to modern quantization theory and microlocal analysis, whereas the second involves methods of C*-algebras and the concepts of superselection sectors and macroscopic observables. In these limits, the classical world does not emerge as a sharply defined objective reality, but rather as an approximate appearance relative to certain "...

  16. Randomness: Quantum versus classical

    Science.gov (United States)

    Khrennikov, Andrei

    2016-05-01

    Recent tremendous development of quantum information theory has led to a number of quantum technological projects, e.g. quantum random generators. This development had stimulated a new wave of interest in quantum foundations. One of the most intriguing problems of quantum foundations is the elaboration of a consistent and commonly accepted interpretation of a quantum state. Closely related problem is the clarification of the notion of quantum randomness and its interrelation with classical randomness. In this short review, we shall discuss basics of classical theory of randomness (which by itself is very complex and characterized by diversity of approaches) and compare it with irreducible quantum randomness. We also discuss briefly “digital philosophy”, its role in physics (classical and quantum) and its coupling to the information interpretation of quantum mechanics (QM).

  17. Kinetic transport in crystals

    CERN Document Server

    Marklof, Jens

    2009-01-01

    One of the central challenges in kinetic theory is the derivation of macroscopic evolution equations--describing, for example, the dynamics of an electron gas--from the underlying fundamental microscopic laws of classical or quantum mechanics. An iconic mathematical model in this research area is the Lorentz gas, which describes an ensemble of non-interacting point particles in an infinite array of spherical scatterers. In the case of a disordered scatterer configuration, the classical results by Gallavotti, Spohn and Boldrighini-Bunimovich-Sinai show that the time evolution of a macroscopic particle cloud is governed, in the limit of small scatterer density (Boltzmann-Grad limit), by the linear Boltzmann equation. In this lecture I will discuss the recent discovery that for a periodic configuration of scatterers the linear Boltzmann equation fails, and the random flight process that emerges in the Boltzmann-Grad limit is substantially more complicated. The key ingredient in the description of the limiting st...

  18. Classical mechanics with Maxima

    CERN Document Server

    Timberlake, Todd Keene

    2016-01-01

    This book guides undergraduate students in the use of Maxima—a computer algebra system—in solving problems in classical mechanics. It functions well as a supplement to a typical classical mechanics textbook. When it comes to problems that are too difficult to solve by hand, computer algebra systems that can perform symbolic mathematical manipulations are a valuable tool. Maxima is particularly attractive in that it is open-source, multiple-platform software that students can download and install free of charge. Lessons learned and capabilities developed using Maxima are easily transferred to other, proprietary software.

  19. Covariantizing Classical Field Theories

    CERN Document Server

    López, Marco Castrillón

    2010-01-01

    We show how to enlarge the covariance group of any classical field theory in such a way that the resulting "covariantized" theory is 'essentially equivalent' to the original. In particular, our technique will render any classical field theory generally covariant, that is, the covariantized theory will be spacetime diffeomorphism-covariant and free of absolute objects. Our results thus generalize the well-known parametrization technique of Dirac and Kucha\\v{r}. Our constructions apply equally well to internal covariance groups, in which context they produce natural derivations of both the Utiyama minimal coupling and St\\"uckelberg tricks.

  20. Problems in classical mechanics

    CERN Document Server

    Katkar, L N

    2014-01-01

    Problems in classical mechanics presents a lucid treatment of the formulations of Lagrangian, Hamiltonian, and the Principles of Calculus of Variations etc. important for the study of modern physics. The study of classical mechanics prepares students to apply the principles and the mathematical tools to solve real life problems. The book also incorporates and discusses in detail topics such as Central Force Motion, Rigid Body Motion and Canonical Transformations. KEY FEATURES: Around 200 solved examples with complete mathematical theory Around 70 examples given as an exercise to test and develop students understanding The physical interpretation of the Hamiltonian is highlighted

  1. Classic Problems of Probability

    CERN Document Server

    Gorroochurn, Prakash

    2012-01-01

    "A great book, one that I will certainly add to my personal library."—Paul J. Nahin, Professor Emeritus of Electrical Engineering, University of New Hampshire Classic Problems of Probability presents a lively account of the most intriguing aspects of statistics. The book features a large collection of more than thirty classic probability problems which have been carefully selected for their interesting history, the way they have shaped the field, and their counterintuitive nature. From Cardano's 1564 Games of Chance to Jacob Bernoulli's 1713 Golden Theorem to Parrondo's 1996 Perplexin

  2. Classical Holographic Codes

    CERN Document Server

    Brehm, Enrico M

    2016-01-01

    In this work, we introduce classical holographic codes. These can be understood as concatenated probabilistic codes and can be represented as networks uniformly covering hyperbolic space. In particular, classical holographic codes can be interpreted as maps from bulk degrees of freedom to boundary degrees of freedom. Interestingly, they are shown to exhibit features similar to those expected from the AdS/CFT correspondence. Among these are a version of the Ryu-Takayanagi formula and intriguing properties regarding bulk reconstruction and boundary representations of bulk operations. We discuss the relation of our findings with expectations from AdS/CFT and, in particular, with recent results from quantum error correction.

  3. Learning Classical Music Club

    CERN Multimedia

    Learning Classical Music Club

    2010-01-01

    There is a new CERN Club called “Learning Classical Music at CERN”. We are aiming to give classical music lessons for different instruments (see link) for students from 5 to 100 years old. We are now ready to start our activities in the CERN barracks. We are now in the enrollment phase and hope to start lessons very soon ! Club info can be found in the list of CERN Club: http://user.web.cern.ch/user/Communication/SocialLifeActivities/Clubs/Clubs.html Salvatore Buontempo Club President

  4. Stabilized enzymes in continuous gas phase reactions

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Fangxiao; LeJeune, K.; Yang, Zhen [Univ. of Pittsburgh, PA (United States)] [and others

    1995-12-01

    We are assessing the utility of enzymes to catalyze reactions in a continuous gas phase reactor. First, alcohol dehydrogenase has been used to oxidize an unsaturated alcohol, 3-methyl-2-buten-1-ol (UOL), to the corresponding unsaturated aldehyde, 3-methyl-2-butenal (UAL). Cofactor NAD{sup +} was regenerated by concomitant acetone reduction to isopropyl alcohol. Second, organophosphorus hydrolase (OPH) has been used to hydrolyze pesticide vapors. In order to control enzyme hydration level, enzyme water adsorption isotherms at different temperature have been studied. Huttig`s isotherm model has been found suitable to describe adsorption behavior. The influence of enzyme hydration level, enzyme loading on glass beads, reaction temperature and flow rate on enzymatic reaction rate and biocatalyst stability were investigated. Reaction kinetics were studied and a kinetic model was proposed. We will also report our attempts to further stabilize enzymes for use in gas reactions by incorporating them into polymer matrices.

  5. Strong Coupling and Classicalization

    CERN Document Server

    Dvali, Gia

    2016-01-01

    Classicalization is a phenomenon in which a theory prevents itself from entering into a strong-coupling regime, by redistributing the energy among many weakly-interacting soft quanta. In this way, the scattering process of some initial hard quanta splits into a large number of soft elementary processes. In short, the theory trades the strong coupling for a high-multiplicity of quanta. At very high energies, the outcome of such a scattering experiment is a production of soft states of high occupation number that are approximately classical. It is evident that black hole creation in particle collision at super-Planckian energies is a result of classicalization, but there is no a priory reason why this phenomenon must be limited to gravity. If the hierarchy problem is solved by classicalization, the LHC has a chance of detecting a tower of new resonances. The lowest-lying resonances must appear right at the strong coupling scale in form of short-lived elementary particles. The heavier members of the tower must b...

  6. Classical Mythology. Fourth Edition.

    Science.gov (United States)

    Morford, Mark P. O.; Lenardon, Robert J.

    Designed for students with little or no background in classical literature, this book introduces the Greek and Roman myths of creation, myths of the gods, Greek sagas and local legends, and presents contemporary theories about the myths. Drawing on Homer, Hesiod, Pindar, Vergil, and others, the book provides many translations and paraphrases of…

  7. Classicism and Romanticism.

    Science.gov (United States)

    Huddleston, Gregory H.

    1993-01-01

    Describes one teacher's methods for introducing to secondary English students the concepts of Classicism and Romanticism in relation to pictures of gardens, architecture, music, and literary works. Outlines how the unit leads to a writing assignment based on collected responses over time. (HB)

  8. Mecanica Clasica (Classical Mechanics)

    OpenAIRE

    Rosu, H. C.

    1999-01-01

    First Internet graduate course on Classical Mechanics in Spanish (Castellano). This is about 80% of the material I covered during the January-June 1999 semester at IFUG in the Mexican city of Leon. English and Romanian versions are in (slow) progress and hopefully will be arXived. For a similar course on Quantum Mechanics, see physics/9808031

  9. Nuclear motion is classical

    CERN Document Server

    Frank, Irmgard

    2016-01-01

    The notion from ab-initio molecular dynamics simulations that nuclear motion is best described by classical Newton dynamics instead of the time-dependent Schr{\\"o}dinger equation is substantiated. In principle a single experiment should bring clarity. Caution is however necessary, as temperature dependent effects must be eliminated when trying to determine the existence of a zero-point energy.

  10. Mecanica Clasica (Classical Mechanics)

    CERN Document Server

    Rosu, H C

    1999-01-01

    First Internet undergraduate course on Classical Mechanics in Spanish (Castellano). This is about 80% of the material I covered during the January-June 1999 semester at IFUG in the Mexican city of Leon. English and Romanian versions are in (slow) progress and hopefully will be arXived. For a similar course on Quantum Mechanics, see physics/9808031

  11. Classical Mechanics Laboratory

    Science.gov (United States)

    Brosing, Juliet W.

    2006-12-01

    At Pacific University we have included a lab with our upper division Classical Mechanics class. We do a combination of physical labs (air resistance, harmonic motion, amusement park physics), Maple labs (software), and projects. Presentation of some of the labs, results and challenges with this course will be included.

  12. Children's Classics. Fifth Edition.

    Science.gov (United States)

    Jordan, Alice M.

    "Children's Classics," a 1947 article by Alice M. Jordan reprinted from "The Horn Book Magazine," examines the dynamics and appeal of some of the most famous books for young readers, including "Alice in Wonderland,""The Wind in the Willows,""Robinson Crusoe," and "Andersen's Fairy Tales." Paul Hein's annotated bibliography, a revision of Jordan's…

  13. Activation of thiamin diphosphate in enzymes.

    Science.gov (United States)

    Hübner, G; Tittmann, K; Killenberg-Jabs, M; Schäffner, J; Spinka, M; Neef, H; Kern, D; Kern, G; Schneider, G; Wikner, C; Ghisla, S

    1998-06-29

    Activation of the coenzyme ThDP was studied by measuring the kinetics of deprotonation at the C2 carbon of thiamin diphosphate in the enzymes pyruvate decarboxylase, transketolase, pyruvate dehydrogenase complex, pyruvate oxidase, in site-specific mutant enzymes and in enzyme complexes containing coenzyme analogues by proton/deuterium exchange detected by 1H-NMR spectroscopy. The respective deprotonation rate constant is above the catalytic constant in all enzymes investigated. The fast deprotonation requires the presence of an activator in pyruvate decarboxylase from yeast, showing the allosteric regulation of this enzyme to be accomplished by an increase in the C2-H dissociation rate of the enzyme-bound thiamin diphosphate. The data of the thiamin diphosphate analogues and of the mutant enzymes show the N1' atom and the 4'-NH2 group to be essential for the activation of the coenzyme and a conserved glutamate involved in the proton abstraction mechanism of the enzyme-bound thiamin diphosphate.

  14. Global Kinetic Analysis of Mammalian E3 Reveals pH-dependent NAD+/NADH Regulation, Physiological Kinetic Reversibility, and Catalytic Optimum.

    Science.gov (United States)

    Moxley, Michael A; Beard, Daniel A; Bazil, Jason N

    2016-02-05

    Mammalian E3 is an essential mitochondrial enzyme responsible for catalyzing the terminal reaction in the oxidative catabolism of several metabolites. E3 is a key regulator of metabolic fuel selection as a component of the pyruvate dehydrogenase complex (PDHc). E3 regulates PDHc activity by altering the affinity of pyruvate dehydrogenase kinase, an inhibitor of the enzyme complex, through changes in reduction and acetylation state of lipoamide moieties set by the NAD(+)/NADH ratio. Thus, an accurate kinetic model of E3 is needed to predict overall mammalian PDHc activity. Here, we have combined numerous literature data sets and new equilibrium spectroscopic experiments with a multitude of independently collected forward and reverse steady-state kinetic assays using pig heart E3. The latter kinetic assays demonstrate a pH-dependent transition of NAD(+) activation to inhibition, shown here, to our knowledge, for the first time in a single consistent data set. Experimental data were analyzed to yield a thermodynamically constrained four-redox-state model of E3 that simulates pH-dependent activation/inhibition and active site redox states for various conditions. The developed model was used to determine substrate/product conditions that give maximal E3 rates and show that, due to non-Michaelis-Menten behavior, the maximal flux is different compared with the classically defined kcat.

  15. Study on discrepancy and correlation of urinary creatinine levels between enzyme method and Jaffe kinetic creatinine assay%酶法与碱性苦味酸法测定尿肌酐值的差异及相关性研究

    Institute of Scientific and Technical Information of China (English)

    史善富; 陶松宁

    2011-01-01

    目的 研究酶法和碱性苦味酸法测定尿肌酐值的差异及相关性,探讨酶法测定肌酐值用于职业接触汞的生物限值的报告.方法 将123名受检者尿液分为两组,分别用0.9%的生理盐水作1∶10及1∶20稀释,用肌氨酸氧化酶法和碱性苦味酸速率法测定两组尿肌酐值,分析两法测定尿肌酐值的差异及相关性.结果 1∶10稀释组尿液肌酐均值苦味酸法低于酶法(P<0.01),相关系数r=0.9955,回归方程Y苦味酸法=0.5726X酶法+392;1∶20稀释组尿液肌酐均值苦味酸法低于酶法(P<0.01),相关系数r=0.9977,回归方程Y苦味酸法=0.5723X酶法+602.结论 两种方法测定的尿液肌酐值差异有统计学意义但具有良好的相关性,可以酶法测定的尿肌酐值计算成苦味酸法测定的尿肌酐值用于职业接触汞的生物限值的报告.%Objective To compare the enzymic method and Jaffe kinetic ereatinine assay for the determination of creatinine in urine and discuss whether using the levels of creatinine by enzymic method to report on biological limits for occupational exposure to mercury. Methods A total of 123 conscript persons were selected as study subjects, they were further divided into two groups, their urine were diluted into 1: 10 and 1: 20 respectively by 0. 9% physiological saline. The levels of urine creatinine were determined by enzymic method and Jaffe kinetic creatinine assay, their differece and correlation were analyzed statistically. Results The creatinine concentrations in urine determined by the Jaffe kinetic creatinine assay were significantly lower than those by enzymic method in 1: 10 group (P < 0.01) and correlation coefficient r = 0.9955, regression equation J'jiiffe kinetic cre.tMm u?a, = 0. 5726 Xmzp^c method + 392; The creatinine concentrations in urine determined by the Jaffe kinetic creatinine assay were significantly lower than those by enzymic method in 1: 20 group ( P < 0. 01) and correlation coefficient r = 0

  16. Biossensor enzimático para detecção de fungicidas ditiocarbamatos: estudo cinético da enzima aldeído desidrogenase e otimização do biossensor Enzymatic biosensor for the detection of dithiocarbamate fungicides: kinetic study of aldehyde dehydrogenase enzyme and biosensor optimization

    Directory of Open Access Journals (Sweden)

    Roberval Soares Lima

    2007-02-01

    Full Text Available Initially, all major factors that affect the rate of the AldH-catalyzed reaction (enzyme concentration, substrate concentration, temperature and pH were investigated. Optimal activity was observed between pH values of 7.5 and 9.5 in the temperature range of 25 to 50 ºC. Kinetic parameters, such as Km (2.92 µmol L-1 and Vmax (1.33 10-2 µmol min-1 demonstrate a strong enzyme-substrate affinity. The sensors were based on screen-printed electrodes modified with the Meldola Blue-Reinecke salt (MBRS combination. Operational conditions (NAD+ and substrate contents, enzyme loading and response time were optimized. Also, two enzyme immobilization procedures were tested: entrapment in poly(vinyl alcohol bearing styrylpyridinium groups (PVA-SbQ and crosslinking with glutaraldehyde. Chronoamperometry was employed to observe the biosensor responses during enzymatic hydrolysis of propionaldehyde and also to construct inhibition curves with maneb and zineb fungicides. Best results were found with the following conditions: [NAD+] = 0.25 mmol L-1; [propionaldehyde] = 80 µmol L-1; enzyme loading = 0.8 U per electrode; response time = 10 min, and inhibition time = 10 min. Current intensities around 103 ± 13 nA with the sensors and good stability was obtained for both immobilization procedures. Detection limits, calculated using 10% inhibition were 31.5 µg L-1 and 35 µg L-1 for maneb and zineb, respectively. Results obtained with other MBRS-modified electrodes consisting of mono and bi-enzymic sensors were compared. The ability to catalyze NADH oxidation by MB was also highlighted.

  17. Food Enzymes

    Science.gov (United States)

    McBroom, Rachel; Oliver-Hoyo, Maria T.

    2007-01-01

    Many students view biology and chemistry as two unrelated, separate sciences; how these courses are generally taught in high schools may do little to change that impression. The study of enzymes provide a great opportunity for both biology and chemistry teachers to share with students the interdisciplinary nature of science. This article describes…

  18. Enzyme immunoassay

    DEFF Research Database (Denmark)

    Feldt-Rasmussen, B; Dinesen, B; Deckert, M

    1985-01-01

    An enzyme linked immunoadsorbent assay for urinary albumin using commercially available reagents is described. The assay range is 2.5-120 micrograms/l. When samples are analysed in two standard dilutions, the assayable albumin concentration range is 2.5-240 mg/l, covering the clinical range from...

  19. Classical Weyl Transverse Gravity

    CERN Document Server

    Oda, Ichiro

    2016-01-01

    We study various classical aspects of the Weyl transverse (WTDiff) gravity in a general space-time dimension. First of all, we clarify a classical equivalence among three kinds of gravitational theories, those are, the conformally-invariant scalar tensor gravity, Einstein's general relativity and the WTDiff gravity via the gauge fixing procedure. Secondly, we show that in the WTDiff gravity the cosmological constant is a mere integration constant as in unimodular gravity, but it does not receive any radiative corrections unlike the unimodular gravity. A key point in this proof is to construct a covariantly conserved energy-momentum tensor, which is achieved on the basis of this equivalence relation. Thirdly, we demonstrate that the Noether current for the Weyl transformation is identically vanishing, thereby implying that the Weyl symmetry existing in both the conformally-invariant scalar tensor gravity and the WTDiff gravity is a "fake" symmetry. We find it possible to extend this proof to all matter fields,...

  20. Classical Diophantine equations

    CERN Document Server

    1993-01-01

    The author had initiated a revision and translation of "Classical Diophantine Equations" prior to his death. Given the rapid advances in transcendence theory and diophantine approximation over recent years, one might fear that the present work, originally published in Russian in 1982, is mostly superseded. That is not so. A certain amount of updating had been prepared by the author himself before his untimely death. Some further revision was prepared by close colleagues. The first seven chapters provide a detailed, virtually exhaustive, discussion of the theory of lower bounds for linear forms in the logarithms of algebraic numbers and its applications to obtaining upper bounds for solutions to the eponymous classical diophantine equations. The detail may seem stark--- the author fears that the reader may react much as does the tourist on first seeing the centre Pompidou; notwithstanding that, Sprind zuk maintainsa pleasant and chatty approach, full of wise and interesting remarks. His emphases well warrant, ...

  1. Electrodynamics classical inconsistencies

    CERN Document Server

    De Souza, M M

    1995-01-01

    The problems of Classical Electrodynamics with the electron equation of motion and with non-integrable singularity of its self-field stress tensor are well known. They are consequences, we show, of neglecting terms that are null off the charge world line but that gives a non null contribution on its world line. The self-field stress tensor of a point classical electron is integrable, there is no causality violation and no conflict with energy conservation in its equation of motion, and there is no need of any kind of renormalization nor of any change in the Maxwell's theory for this. (This is part of the paper hep-th/9510160, stripped , for simplicity, of its non-Minkowskian geometrization of causality and of its discussion about the physical meaning of the Maxwell-Faraday concept of field).

  2. Injuries in classical ballet

    Directory of Open Access Journals (Sweden)

    Adriana Coutinho de Azevedo Guimarães

    2008-06-01

    Full Text Available This study aimed to elucidate what injuries are most likely to occur due to classical ballet practice. The research used national and international bibliography. The bibliography analysis indicated that technical and esthetical demands lead to a practice of non-anatomical movements, causing the ballet dancer to suffer from a number of associated lesions. Most of the injuries are caused by technical mistakes and wrong training. Troubles in children are usually due to trying to force external rotation at hip level and to undue use of point ballet slippers. The commonest lesions are in feet and ankles, followed by knees and hips. The rarest ones are in the upper limbs. These injuries are caused by exercise excess, by repetitions always in the same side and by wrong and early use of point slippers. The study reached the conclusion that incorrect application of classical ballet technique predisposes the dancers to characteristic injuries.

  3. Classical and statistical thermodynamics

    CERN Document Server

    Rizk, Hanna A

    2016-01-01

    This is a text book of thermodynamics for the student who seeks thorough training in science or engineering. Systematic and thorough treatment of the fundamental principles rather than presenting the large mass of facts has been stressed. The book includes some of the historical and humanistic background of thermodynamics, but without affecting the continuity of the analytical treatment. For a clearer and more profound understanding of thermodynamics this book is highly recommended. In this respect, the author believes that a sound grounding in classical thermodynamics is an essential prerequisite for the understanding of statistical thermodynamics. Such a book comprising the two wide branches of thermodynamics is in fact unprecedented. Being a written work dealing systematically with the two main branches of thermodynamics, namely classical thermodynamics and statistical thermodynamics, together with some important indexes under only one cover, this treatise is so eminently useful.

  4. Randomness: quantum versus classical

    CERN Document Server

    Khrennikov, Andrei

    2015-01-01

    Recent tremendous development of quantum information theory led to a number of quantum technological projects, e.g., quantum random generators. This development stimulates a new wave of interest in quantum foundations. One of the most intriguing problems of quantum foundations is elaboration of a consistent and commonly accepted interpretation of quantum state. Closely related problem is clarification of the notion of quantum randomness and its interrelation with classical randomness. In this short review we shall discuss basics of classical theory of randomness (which by itself is very complex and characterized by diversity of approaches) and compare it with irreducible quantum randomness. The second part of this review is devoted to the information interpretation of quantum mechanics (QM) in the spirit of Zeilinger and Brukner (and QBism of Fuchs et al.) and physics in general (e.g., Wheeler's "it from bit") as well as digital philosophy of Chaitin (with historical coupling to ideas of Leibnitz). Finally, w...

  5. Computation in Classical Mechanics

    CERN Document Server

    Timberlake, Todd

    2007-01-01

    There is a growing consensus that physics majors need to learn computational skills, but many departments are still devoid of computation in their physics curriculum. Some departments may lack the resources or commitment to create a dedicated course or program in computational physics. One way around this difficulty is to include computation in a standard upper-level physics course. An intermediate classical mechanics course is particularly well suited for including computation. We discuss the ways we have used computation in our classical mechanics courses, focusing on how computational work can improve students' understanding of physics as well as their computational skills. We present examples of computational problems that serve these two purposes. In addition, we provide information about resources for instructors who would like to include computation in their courses.

  6. Kinetic Interface

    DEFF Research Database (Denmark)

    2009-01-01

    A kinetic interface for orientation detection in a video training system is disclosed. The interface includes a balance platform instrumented with inertial motion sensors. The interface engages a participant's sense of balance in training exercises.......A kinetic interface for orientation detection in a video training system is disclosed. The interface includes a balance platform instrumented with inertial motion sensors. The interface engages a participant's sense of balance in training exercises....

  7. Lectures on classical electrodynamics

    CERN Document Server

    Englert, Berthold-Georg

    2014-01-01

    These lecture notes cover classical electrodynamics at the level of advanced undergraduates or postgraduates. There is a strong emphasis on the general features of the electromagnetic field and, in particular, on the properties of electromagnetic radiation. It offers a comprehensive and detailed, as well as self-contained, account of material that can be covered in a one-semester course for students with a solid undergraduate knowledge of basic electricity and magnetism.

  8. Concepts of classical optics

    CERN Document Server

    Strong, John

    2004-01-01

    An intermediate course in optics, this volume explores both experimental and theoretical concepts, offering practical knowledge of geometrical optics that will enhance students' comprehension of any relevant applied science. Its exposition of the concepts of classical optics is presented with a minimum of mathematical detail but presumes some knowledge of calculus, vectors, and complex numbers.Subjects include light as wave motion; superposition of wave motions; electromagnetic waves; interaction of light and matter; velocities and scattering of light; polarized light and dielectric boundarie

  9. Revisiting a Classic

    Science.gov (United States)

    Rogers, Ibram

    2008-01-01

    As a 26-year-old English teacher in 1958, Chinua Achebe had no idea that the book he was writing would become a literary classic, not only in Africa but also throughout the world. He could only try to articulate the feelings he had for his countrymen and women. Achebe had a burning desire to tell the true story of Africa and African humanity. The…

  10. Optimization of Inhibitory Reaction System of Folium Mori Extracts on Alpha-glucosidase Activity and Enzyme Kinetics%桑叶提取物抑制α-葡萄糖苷酶活性体系的优化及动力学研究

    Institute of Scientific and Technical Information of China (English)

    吕欢; 罗明琍; 方飞; 吴新荣

    2012-01-01

    Objective To establish an optimized inhibitory reaction system for the research of Folium Mori extracts in inhibiting alpha-glucosidase activity and to investigate the enzyme kinetics. Methods We studied the inhibitive effects of Folium Mori extracts on alpha-glucosidase by using enzyme-inhibitor model and optimized the reaction system by optimizing the substrate concentration, the amount of enzyme and the reaction time. We also investigated the inhibitory kinetic characteristics of extracts by using Lineweaver-Burk method. Results The optimized reaction system consisted of 40 mg/mL of the substrate concentration, 0.2 mL of enzyme and 25 min for reaction time. Inhibitory kinetic results indicated that the extracts were mixed competitive inhibitors, but the positive drug was noncompetitive inhibitor. Conclusion Folium Mori extracts have inhibitory effect on a- glucosidase activity, and the inhibitory type is in mixed competitive inhibition, which are expected to be used to develop as anti-diabetes drugs.%目的 研究桑叶提取物抑制α-葡萄糖苷酶活性反应体系的条件优化及酶动力学.方法 采用酶-抑制剂模型法,优化底物浓度、酶用量和反应时间,确定最佳反应体系,研究桑叶提取物对α-葡萄糖苷酶的抑制作用,并采用双倒数作图法研究桑叶提取物的酶抑制动力学特性.结果 当蔗糖浓度为40 mg/mL,酶用量为0.2mL(约1.6 U/mL),反应时间为25 min时为最佳反应体系;酶抑制动力学实验表明,桑叶提取物的抑制类型为混合型竞争性抑制,阳性对照为非竞争性抑制.结论 桑叶提取物对α-葡萄糖苷酶活性有抑制作用;桑叶提取物对α-葡萄糖苷酶抑制活性为混合性竞争性抑制,可以用于开发降糖药物.

  11. Classical Trace Anomaly

    OpenAIRE

    Farhoudi, M.

    1995-01-01

    We seek an analogy of the mathematical form of the alternative form of Einstein's field equations for Lovelock's field equations. We find that the price for this analogy is to accept the existence of the trace anomaly of the energy-momentum tensor even in classical treatments. As an example, we take this analogy to any generic second order Lagrangian and exactly derive the trace anomaly relation suggested by Duff. This indicates that an intrinsic reason for the existence of such a relation sh...

  12. The classical task

    DEFF Research Database (Denmark)

    Gillis, Steven; Souman, Agnita; Dhollander, Sim

    -year-olds to 9-year-olds. The experiment was also administered with a control group of adults. Procedure: The procedure consists of a classical set-up in which the subjects are shown pictures of objects. On presenting each object, the test leader says: "Here is a X (name of the object)". The next...... articulated: Prediction 1: a global analysis of the plural forms provided by the subjects is expected to show an increase of the correct responses as children grow older. Prediction 2: As to suffix selection, we expect that the plural of nouns selecting a fully predictable suffix will be more readily mastered...

  13. A Classic Through Eternity

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    FIVE years ago, an ancient Chinese air was beamed to outer space as a PR exercise. To humankind, music is a universal language, so the tune seemed an ideal medium for communication with extraterrestrial intelligence. So far there has been no response, but it is believed that the tune will play for a billion years, and eventually be heard and understood. The melody is called High Mountain and Flowing Stream, and it is played on the guqin, a seven-stringed classical musical instrument similar to the zither.

  14. On Classical Ideal Gases

    Directory of Open Access Journals (Sweden)

    Laurent Chusseau

    2013-02-01

    Full Text Available We show that the thermodynamics of ideal gases may be derived solely from the Democritean concept of corpuscles moving in vacuum plus a principle of simplicity, namely that these laws are independent of the laws of motion, aside from the law of energy conservation. Only a single corpuscle in contact with a heat bath submitted to a z and t-invariant force is considered. Most of the end results are known but the method appears to be novel. The mathematics being elementary, the present paper should facilitate the understanding of the ideal gas law and of classical thermodynamics even though not-usually-taught concepts are being introduced.

  15. Semi-classical Electrodynamics

    Science.gov (United States)

    Lestone, John

    2016-03-01

    Quantum electrodynamics is complex and its associated mathematics can appear overwhelming for those not trained in this field. We describe semi-classical approaches that can be used to obtain a more intuitive physical feel for several QED processes including electro-statics, Compton scattering, pair annihilation, the anomalous magnetic moment, and the Lamb shift, that could be taught easily to undergraduate students. Any physicist who brings their laptop to the talk will be able to build spread sheets in less than 10 minutes to calculate g/2 =1.001160 and a Lamb shift of 1057 MHz.

  16. Classical cytogenetics: karyotyping techniques.

    Science.gov (United States)

    Bates, Steven E

    2011-01-01

    Classical cytogenetics by karyotyping has been utilized in clinical research laboratories for more than 50 years and remains the key method used in the stem cell laboratory to assess the genetic stability of stem cell cultures. It is currently the most readily accessible method for detecting chromosomal abnormalities in pluripotent stem cell cultures. This chapter will describe (1) how to prepare a culture to maximize the number of metaphase cells, (2) how to prepare slides containing chromosome spreads (3) methods used to stain chromosomes, and (4) how to interpret the cytogenetic report.

  17. Mechanics classical and quantum

    CERN Document Server

    Taylor, T T

    2015-01-01

    Mechanics: Classical and Quantum explains the principles of quantum mechanics via the medium of analytical mechanics. The book describes Schrodinger's formulation, the Hamilton-Jacobi equation, and the Lagrangian formulation. The author discusses the Harmonic Oscillator, the generalized coordinates, velocities, as well as the application of the Lagrangian formulation to systems that are partially or entirely electromagnetic in character under certain conditions. The book examines waves on a string under tension, the isothermal cavity radiation, and the Rayleigh-Jeans result pertaining to the e

  18. Kinetics and

    Directory of Open Access Journals (Sweden)

    Mojtaba Ahmadi

    2016-11-01

    Full Text Available The aqueous degradation of Reactive Yellow 84 (RY84 by potassium peroxydisulfate (K2S2O8 has been studied in laboratory scale experiments. The effect of the initial concentrations of potassium peroxydisulfate and RY84, pH and temperature on RY84 degradation were also examined. Experimental data were analyzed using first and second-order kinetics. The degradation kinetics of RY84 of the potassium peroxydisulfate process followed the second-order reaction kinetics. These rate constants have an extreme values similar to of 9.493 mM−1min−1 at a peroxydisulfate dose of 4 mmol/L. Thermodynamic parameters such as activation (Ea and Gibbs free energy (ΔG° were also evaluated. The negative value of ΔGo and Ea shows the spontaneous reaction natural conditions and exothermic nature.

  19. Probability representation of classical states

    NARCIS (Netherlands)

    Man'ko, OV; Man'ko, [No Value; Pilyavets, OV

    2005-01-01

    Probability representation of classical states described by symplectic tomograms is discussed. Tomographic symbols of classical observables which are functions on phase-space are studied. Explicit form of kernel of commutative star-product of the tomographic symbols is obtained.

  20. Cloning and expression of the liver and muscle isoforms of ovine carnitine palmitoyltransferase 1 : residues within the N-terminus of the muscle isoform influence the kinetic properties of the enzyme

    NARCIS (Netherlands)

    Price, NT; Jackson, VN; van der Leij, FR; Cameron, JM; Travers, MT; Bartelds, B; Huijkman, NC; Zammit, VA

    2003-01-01

    Fatty acid and ketone body metabolism differ considerably between monogastric and ruminant species. The regulation of the key enzymes involved may differ accordingly. Carnitine palmitoyltransferase 1 (CPT 1) is the key locus for the control of long-chain fatty acid P-oxidation and liver ketogenesis.

  1. Classical Trajectories and Quantum Spectra

    Science.gov (United States)

    Mielnik, Bogdan; Reyes, Marco A.

    1996-01-01

    A classical model of the Schrodinger's wave packet is considered. The problem of finding the energy levels corresponds to a classical manipulation game. It leads to an approximate but non-perturbative method of finding the eigenvalues, exploring the bifurcations of classical trajectories. The role of squeezing turns out decisive in the generation of the discrete spectra.

  2. Kinetics of reactions of the Actinomadura R39 DD-peptidase with specific substrates.

    Science.gov (United States)

    Adediran, S A; Kumar, Ish; Nagarajan, Rajesh; Sauvage, Eric; Pratt, R F

    2011-01-25

    The Actinomadura R39 DD-peptidase catalyzes the hydrolysis and aminolysis of a number of small peptides and depsipeptides. Details of its substrate specificity and the nature of its in vivo substrate are not, however, well understood. This paper describes the interactions of the R39 enzyme with two peptidoglycan-mimetic substrates 3-(D-cysteinyl)propanoyl-D-alanyl-D-alanine and 3-(D-cysteinyl)propanoyl-D-alanyl-D-thiolactate. A detailed study of the reactions of the former substrate, catalyzed by the enzyme, showed DD-carboxypeptidase, DD-transpeptidase, and DD-endopeptidase activities. These results confirm the specificity of the enzyme for a free D-amino acid at the N-terminus of good substrates and indicated a preference for extended D-amino acid leaving groups. The latter was supported by determination of the structural specificity of amine nucleophiles for the acyl-enzyme generated by reaction of the enzyme with the thiolactate substrate. It was concluded that a specific substrate for this enzyme, and possibly the in vivo substrate, may consist of a partly cross-linked peptidoglycan polymer where a free side chain N-terminal un-cross-linked amino acid serves as the specific acyl group in an endopeptidase reaction. The enzyme is most likely a DD-endopeptidase in vivo. pH-rate profiles for reactions of the enzyme with peptides, the thiolactate named above, and β-lactams indicated the presence of complex proton dissociation pathways with sticky substrates and/or protons. The local structure of the active site may differ significantly for reactions of peptides and β-lactams. Solvent kinetic deuterium isotope effects indicate the presence of classical general acid/base catalysis in both acylation and deacylation; there is no evidence of the low fractionation factor active site hydrogen found previously in class A and C β-lactamases.

  3. Perspective: Quantum or classical coherence?

    Science.gov (United States)

    Miller, William H

    2012-06-07

    Some coherence effects in chemical dynamics are described correctly by classical mechanics, while others only appear in a quantum treatment--and when these are observed experimentally it is not always immediately obvious whether their origin is classical or quantum. Semiclassical theory provides a systematic way of adding quantum coherence to classical molecular dynamics and thus provides a useful way to distinguish between classical and quantum coherence. Several examples are discussed which illustrate both cases. Particularly interesting is the situation with electronically non-adiabatic processes, where sometimes whether the coherence effects are classical or quantum depends on what specific aspects of the process are observed.

  4. Mechanical Systems, Classical Models

    CERN Document Server

    Teodorescu, Petre P

    2009-01-01

    This third volume completes the Work Mechanical Systems, Classical Models. The first two volumes dealt with particle dynamics and with discrete and continuous mechanical systems. The present volume studies analytical mechanics. Topics like Lagrangian and Hamiltonian mechanics, the Hamilton-Jacobi method, and a study of systems with separate variables are thoroughly discussed. Also included are variational principles and canonical transformations, integral invariants and exterior differential calculus, and particular attention is given to non-holonomic mechanical systems. The author explains in detail all important aspects of the science of mechanics, regarded as a natural science, and shows how they are useful in understanding important natural phenomena and solving problems of interest in applied and engineering sciences. Professor Teodorescu has spent more than fifty years as a Professor of Mechanics at the University of Bucharest and this book relies on the extensive literature on the subject as well as th...

  5. Grassmannization of classical models

    CERN Document Server

    Pollet, Lode; Prokof'ev, Nikolay V; Svistunov, Boris V

    2016-01-01

    Applying Feynman diagrammatics to non-fermionic strongly correlated models with local constraints might seem generically impossible for two separate reasons: (i) the necessity to have a Gaussian (non-interacting) limit on top of which the perturbative diagrammatic expansion is generated by Wick's theorem, and (ii) the Dyson's collapse argument implying that the expansion in powers of coupling constant is divergent. We show that for arbitrary classical lattice models both problems can be solved/circumvented by reformulating the high-temperature expansion (more generally, any discrete representation of the model) in terms of Grassmann integrals. Discrete variables residing on either links, plaquettes, or sites of the lattice are associated with the Grassmann variables in such a way that the partition function (and correlations) of the original system and its Grassmann-field counterpart are identical. The expansion of the latter around its Gaussian point generates Feynman diagrams. A proof-of-principle implement...

  6. Classical and quantum cosmology

    CERN Document Server

    Calcagni, Gianluca

    2017-01-01

    This comprehensive textbook is devoted to classical and quantum cosmology, with particular emphasis on modern approaches to quantum gravity and string theory and on their observational imprint. It covers major challenges in theoretical physics such as the big bang and the cosmological constant problem. An extensive review of standard cosmology, the cosmic microwave background, inflation and dark energy sets the scene for the phenomenological application of all the main quantum-gravity and string-theory models of cosmology. Born of the author's teaching experience and commitment to bridging the gap between cosmologists and theoreticians working beyond the established laws of particle physics and general relativity, this is a unique text where quantum-gravity approaches and string theory are treated on an equal footing. As well as introducing cosmology to undergraduate and graduate students with its pedagogical presentation and the help of 45 solved exercises, this book, which includes an ambitious bibliography...

  7. Citation classics in epilepsy

    Directory of Open Access Journals (Sweden)

    Maryann Wilson

    2013-01-01

    Full Text Available BACKGROUND: The impact of a scientific article is proportional to the citations it has received. In this study, we set out to identify the most cited works in epileptology in order to evaluate research trends in this field. METHODS: According to the Web of Science database, articles with more than 400 citations qualify as "citation classics". We conducted a literature search on the ISI Web of Science bibliometric database for scientific articles relevant to epilepsy. RESULTS: We retrieved 67 highly cited articles (400 or more citations, which were published in 31 journals: 17 clinical studies, 42 laboratory studies, 5 reviews and 3 classification articles. Clinical studies consisted of epidemiological analyses (n=3, studies on the clinical phenomenology of epilepsy (n=5 – including behavioral and prognostic aspects – and articles focusing on pharmacological (n=6 and non-pharmacological (n=3 treatment. The laboratory studies dealt with genetics (n=6, animal models (n=27, and neurobiology (n=9 – including both neurophysiology and neuropathology studies. The majority (61% of citation classics on epilepsy were published after 1986, possibly reflecting the expansion of research interest in laboratory studies driven by the development of new methodologies, specifically in the fields of genetics and animal models. Consequently, clinical studies were highly cited both before and after the mid 80s, whilst laboratory researches became widely cited after 1990. CONCLUSIONS: Our study indicates that the main drivers of scientific impact in the field of epileptology have increasingly become genetic and neurobiological studies, along with research on animal models of epilepsy. These articles are able to gain the highest numbers of citations in the time span of a few years and suggest potential directions for future research.

  8. Generalization of the Activated Complex Theory of Reaction Rates. II. Classical Mechanical Treatment

    Science.gov (United States)

    Marcus, R. A.

    1964-01-01

    In its usual classical form activated complex theory assumes a particular expression for the kinetic energy of the reacting system -- one associated with a rectilinear motion along the reaction coordinate. The derivation of the rate expression given in the present paper is based on the general kinetic energy expression.

  9. Cinética de Inactivación de la Enzima Peroxidasa, Color y Textura en Papa Criolla (Solanum tuberosum Grupo phureja sometida a tres Condiciones de Escaldado Kinetics of Peroxidase Enzyme Inactivation, Color and Texture in Golden Potato (Solanum tuberosum phureja group under three Blanching Conditions

    Directory of Open Access Journals (Sweden)

    Rolando Mendoza

    2012-01-01

    Full Text Available El objetivo de este trabajo fue determinar la cinética de inactivación de la enzima peroxidasa y la cinética del color y textura en tubérculos de papa criolla (Solanum tuberosum grupo phureja sometidos a escaldado mediante agua a 80°C, 90°C y vapor saturado a 93°C. Los coeficientes de transferencia de calor para el escaldado en agua oscilaron entre 214 y 230 W °C-1 m-2 mientras que para vapor saturado fueron en promedio 84.5 W °C-1 m-2. A diferentes tiempos de precocción el tubérculo de papa criolla presentó una zona en donde ocurrió pardeamiento enzimático, hecho que fue modelado según la cinética de Michaelis-Menten y otra zona en donde el cambio de color disminuyó debido a la inactivación enzimática que correspondió a una cinética de primer orden, de manera similar al comportamiento de la textura. La inactivación de la enzima peroxidasa obedeció al mecanismo de Lumry-Eyring.The objective of this work was to determine the inactivation kinetics of peroxidase enzyme and the change in color and texture in golden potato tubers (Solanum tuberosum phureja group exposed to blanching with water at 80°C, 90°C and to saturated steam at 93°C. The heat transfer coefficients for the blanch water ranged between 214 and 230 W °C-1 m-2 while for saturated steam they showed an average of 84.5 W °C-1 m-2. At different pre-cooking times the golden potato tuber showed an area that developed enzymatic browning, which was fitted to the Michaelis-Menten kinetic model and another area where the color change decreased due to enzyme inactivation, which corresponded to first order kinetics, in a similar way as the texture behavior. The inactivation of peroxidase enzyme followed the Lumry-Eyring mechanism.

  10. Supramolecular Tectonics for Enzyme-like Reagents

    Institute of Scientific and Technical Information of China (English)

    MAO; LuYuan

    2001-01-01

    The enzyme-likes and bioactive species were closely related with the life phenomena and served as the reagent of bioassy1,2. In present works, the flow cytometry (FCM) and rapid-scanning stopped-flow (RSSF) spectroscopy combine with the stopped-flow difference UV/Vis spectra, FT-IR and other methods of assay, being used to study the biomimetic reaction and enzyme mimic. Based on catalytic kinetics of enzyme reaction3,4, the reaction mechanisms of the enzyme-likes had been studied and some new methods of kinetic determination were proposed. The study and methods not only provided the basic theoretical models for the life science, but also widened the application fields of biomimetic and analytical chemistry. The main contents of our works and the supramolecular models can be described as follows:  ……

  11. Enzyme Reactions in Nanoporous, Picoliter Volume Containers

    Energy Technology Data Exchange (ETDEWEB)

    Siuti, Piro [ORNL; Retterer, Scott T [ORNL; Choi, Chang Kyoung [Michigan Technological University; Doktycz, Mitchel John [ORNL

    2012-01-01

    Advancements in nanoscale fabrication allow creation of small volume reaction containers that can facilitate the screening and characterization of enzymes. A porous, ~19 pL volume vessel has been used in this work to carry out enzyme reactions under varying substrate concentrations. Glucose oxidase and horseradish peroxidase can be contained in these structures and diffusively fed with a solution containing glucose and the fluorogenic substrate Amplex Red through the engineered nanoscale pore structure. Fluorescent microscopy was used to monitor the reaction, which was carried out under microfluidic control. Kinetic characteristics of the enzyme were evaluated and compared with results from conventional scale reactions. These picoliter, nanoporous containers can facilitate quick determination of enzyme kinetics in microfluidic systems without the requirement of surface tethering and can be used for applications in drug discovery, clinical diagnostics and high-throughput screening.

  12. Protein engineering of enzymes for process applications

    DEFF Research Database (Denmark)

    Woodley, John M

    2013-01-01

    Scientific progress in the field of enzyme modification today enables the opportunity to tune a given biocatalyst for a specific industrial application. Much work has been focused on extending the substrate repertoire and altering selectivity. Nevertheless, it is clear that many new forthcoming...... opportunities will be targeted on modification to enable process application. This article discusses the challenges involved in enzyme modification focused on process requirements, such as the need to fulfill reaction thermodynamics, specific activity under the required conditions, kinetics at required...

  13. Kinetic Biochemistry

    Directory of Open Access Journals (Sweden)

    Lorentz JÄNTSCHI

    2003-03-01

    Full Text Available Mathematics and computer programming have a major contribution to chemistry. Two directions can be identified: one that searches and tries (rich to explain the structural binding and shape of the chemical compounds [1] with major applications in QSPR/QSAR studies [2], and applied sciences such as engineering of materials or agriculture [3]; the second direction is to models the kinetic processes that are involved in chemical reactions [4]. Many such models are available here. The present paper describes three variants of well the known kinetic models and presents the mathematical equations associated with them. The differential equations are numerically solved and fitted with MathCad program. [1] Diudea M., Gutman I., Jäntschi L., Molecular Topology, Nova Science, Huntington, New York, 332 p., 2001, 2002. [2] Diudea M. V., Ed., QSPR / QSAR Studies by Molecular Descriptors, Nova Science, Huntington, New York, 438 p., 2001. [3] Jäntschi L., Microbiology and Toxicology. Phytochemistry Studies (in Romanian, Amici, Cluj-Napoca, 184 p., 2003. [4] Jäntschi L., Unguresan M., Physical Chemistry. Molecular Kinetic and Dynamic (in Romanian, Mediamira, Cluj-Napoca, 159 p., 2001.

  14. Physisorption kinetics

    CERN Document Server

    Kreuzer, Hans Jürgen

    1986-01-01

    This monograph deals with the kinetics of adsorption and desorption of molecules physisorbed on solid surfaces. Although frequent and detailed reference is made to experiment, it is mainly concerned with the theory of the subject. In this, we have attempted to present a unified picture based on the master equation approach. Physisorption kinetics is by no means a closed and mature subject; rather, in writing this monograph we intended to survey a field very much in flux, to assess its achievements so far, and to give a reasonable basis from which further developments can take off. For this reason we have included many papers in the bibliography that are not referred to in the text but are of relevance to physisorption. To keep this monograph to a reasonable size, and also to allow for some unity in the presentation of the material, we had to omit a number of topics related to physisorption kinetics. We have not covered to any extent the equilibrium properties of physisorbed layers such as structures, phase tr...

  15. Classical understanding of electron vortex beams in a uniform magnetic field

    Science.gov (United States)

    Han, Yeong Deok; Choi, Taeseung

    2017-04-01

    Recently, interesting observations on electron vortex beams have been made. We propose a classical model that shows vortex-like motion due to suitably-synchronized motion of each electron's cyclotron motion in a uniform magnetic field. It is shown that some basic features of electron vortex beams in a uniform magnetic field, such as azimuthal currents, the relation between energy and kinetic angular momentum, and the parallel-axis theorem are understandable by using this classical model. We also show that the time-dependence of kinetic angular momentum of electron vortex beams could be understood as an effect of a specific nonuniform distribution of classical electrons.

  16. Highly efficient enzyme encapsulation in a protein nanocage: towards enzyme catalysis in a cellular nanocompartment mimic

    Science.gov (United States)

    Schoonen, Lise; Nolte, Roeland J. M.; van Hest, Jan C. M.

    2016-07-01

    The study of enzyme behavior in small nanocompartments is crucial for the understanding of biocatalytic processes in the cellular environment. We have developed an enzymatic conjugation strategy to attach a model enzyme to the interior of a cowpea chlorotic mottle virus capsid. It is shown that with this methodology high encapsulation efficiencies can be achieved. Additionally, we demonstrate that the encapsulation does not affect the enzyme performance in terms of a decreased activity or a hampered substrate diffusion. Finally, it is shown that the encapsulated enzymes are protected against proteases. We believe that our strategy can be used to study enzyme kinetics in an environment that approaches physiological conditions.The study of enzyme behavior in small nanocompartments is crucial for the understanding of biocatalytic processes in the cellular environment. We have developed an enzymatic conjugation strategy to attach a model enzyme to the interior of a cowpea chlorotic mottle virus capsid. It is shown that with this methodology high encapsulation efficiencies can be achieved. Additionally, we demonstrate that the encapsulation does not affect the enzyme performance in terms of a decreased activity or a hampered substrate diffusion. Finally, it is shown that the encapsulated enzymes are protected against proteases. We believe that our strategy can be used to study enzyme kinetics in an environment that approaches physiological conditions. Electronic supplementary information (ESI) available: Experimental procedures for the cloning, expression, and purification of all proteins, as well as supplementary figures and calculations. See DOI: 10.1039/c6nr04181g

  17. Classical Fourier analysis

    CERN Document Server

    Grafakos, Loukas

    2014-01-01

    The main goal of this text is to present the theoretical foundation of the field of Fourier analysis on Euclidean spaces. It covers classical topics such as interpolation, Fourier series, the Fourier transform, maximal functions, singular integrals, and Littlewood–Paley theory. The primary readership is intended to be graduate students in mathematics with the prerequisite including satisfactory completion of courses in real and complex variables. The coverage of topics and exposition style are designed to leave no gaps in understanding and stimulate further study. This third edition includes new Sections 3.5, 4.4, 4.5 as well as a new chapter on “Weighted Inequalities,” which has been moved from GTM 250, 2nd Edition. Appendices I and B.9 are also new to this edition.  Countless corrections and improvements have been made to the material from the second edition. Additions and improvements include: more examples and applications, new and more relevant hints for the existing exercises, new exercises, and...

  18. Extended symmetrical classical electrodynamics.

    Science.gov (United States)

    Fedorov, A V; Kalashnikov, E G

    2008-03-01

    In this paper, we discuss a modification of classical electrodynamics in which "ordinary" point charges are absent. The modified equations contain additional terms describing the induced charges and currents. The densities of the induced charges and currents depend on the vector k and the vectors of the electromagnetic field, E and B . It is shown that the vectors E and B can be defined in terms of two four-potentials and the components of k are the components of a four-tensor of the third rank. The Lagrangian of the modified electrodynamics is defined. The conditions are derived at which only one four-potential determines the behavior of the electromagnetic field. It is also shown that static modified electrodynamics can describe the electromagnetic field in the inner region of an electric monopole. In the outer region of the electric monopole the electric field is governed by the Maxwell equations. It follows from boundary conditions at the interface between the inner and outer regions of the monopole that the vector k has a discrete spectrum. The electric and magnetic fields, energy, and angular momentum of the monopole are found for different eigenvalues of k .

  19. Grassmannization of classical models

    Science.gov (United States)

    Pollet, Lode; Kiselev, Mikhail N.; Prokof'ev, Nikolay V.; Svistunov, Boris V.

    2016-11-01

    Applying Feynman diagrammatics to non-fermionic strongly correlated models with local constraints might seem generically impossible for two separate reasons: (i) the necessity to have a Gaussian (non-interacting) limit on top of which the perturbative diagrammatic expansion is generated by Wick’s theorem, and (ii) Dyson’s collapse argument implying that the expansion in powers of coupling constant is divergent. We show that for arbitrary classical lattice models both problems can be solved/circumvented by reformulating the high-temperature expansion (more generally, any discrete representation of the model) in terms of Grassmann integrals. Discrete variables residing on either links, plaquettes, or sites of the lattice are associated with the Grassmann variables in such a way that the partition function (as well as all correlation functions) of the original system and its Grassmann-field counterpart are identical. The expansion of the latter around its Gaussian point generates Feynman diagrams. Our work paves the way for studying lattice gauge theories by treating bosonic and fermionic degrees of freedom on equal footing.

  20. Time, classical and quantum

    Science.gov (United States)

    Aniello, P.; Ciaglia, F. M.; Di Cosmo, F.; Marmo, G.; Pérez-Pardo, J. M.

    2016-10-01

    We propose a new point of view regarding the problem of time in quantum mechanics, based on the idea of replacing the usual time operator T with a suitable real-valued function T on the space of physical states. The proper characterization of the function T relies on a particular relation with the dynamical evolution of the system rather than with the infinitesimal generator of the dynamics (Hamiltonian). We first consider the case of classical hamiltonian mechanics, where observables are functions on phase space and the tools of differential geometry can be applied. The idea is then extended to the case of the unitary evolution of pure states of finite-level quantum systems by means of the geometric formulation of quantum mechanics. It is found that T is a function on the space of pure states which is not associated with any self-adjoint operator. The link between T and the dynamical evolution is interpreted as defining a simultaneity relation for the states of the system with respect to the dynamical evolution itself. It turns out that different dynamical evolutions lead to different notions of simultaneity, i.e., the notion of simultaneity is a dynamical notion.

  1. Classical competing risks

    CERN Document Server

    Crowder, Martin J

    2001-01-01

    If something can fail, it can often fail in one of several ways and sometimes in more than one way at a time. There is always some cause of failure, and almost always, more than one possible cause. In one sense, then, survival analysis is a lost cause. The methods of Competing Risks have often been neglected in the survival analysis literature. Written by a leading statistician, Classical Competing Risks thoroughly examines the probability framework and statistical analysis of data of Competing Risks. The author explores both the theory of the subject and the practicalities of fitting the models to data. In a coherent, self-contained, and sequential account, the treatment moves from the bare bones of the Competing Risks setup and the associated likelihood functions through survival analysis using hazard functions. It examines discrete failure times and the difficulties of identifiability, and concludes with an introduction to the counting-process approach and the associated martingale theory.With a dearth of ...

  2. Classics in radio astronomy

    CERN Document Server

    Sullivan, Woodruff Turner

    1982-01-01

    Radio techniques were the nrst to lead astronomy away from the quiescent and limited Universe revealed by traditional observations at optical wave­ lengths. In the earliest days of radio astronomy, a handful of radio physicists and engineers made one startling discovery after another as they opened up the radio sky. With this collection of classic papers and the extensive intro­ ductory material, the reader can experience these exciting discoveries, as well as understand the developing techniques and follow the motivations which prompted the various lines of inquiry. For instance he or she will follow in detail the several attempts to detect radio waves from the sun at the turn of the century; the unravelling by Jansky of a "steady hiss type static"; the incredible story of Reber who built a 9 meter dish in his backyard in 1937 and then mapped the Milky Way; the vital discoveries by Hey and colleagues of radio bursts from the Sun and of a discrete source in the constellation of Cygnus; the development of re...

  3. A robust methodology for kinetic model parameter estimation for biocatalytic reactions

    DEFF Research Database (Denmark)

    Al-Haque, Naweed; Andrade Santacoloma, Paloma de Gracia; Lima Afonso Neto, Watson;

    2012-01-01

    Effective estimation of parameters in biocatalytic reaction kinetic expressions are very important when building process models to enable evaluation of process technology options and alternative biocatalysts. The kinetic models used to describe enzyme-catalyzed reactions generally include several...

  4. Kinetic Transport in Crystals

    Science.gov (United States)

    Marklof, Jens

    2010-03-01

    One of the central challenges in kinetic theory is the derivation of macroscopic evolution equations--describing, for example, the dynamics of an electron gas--from the underlying fundamental microscopic laws of classical or quantum mechanics. An iconic mathematical model in this research area is the Lorentz gas, which describes an ensemble of non-interacting point particles in an infinite array of spherical scatterers. In the case of a disordered scatterer configuration, the classical results by Gallavotti, Spohn and Boldrighini-Bunimovich-Sinai show that the time evolution of a macroscopic particle cloud is governed, in the limit of small scatterer density (Boltzmann-Grad limit), by the linear Boltzmann equation. In this lecture I will discuss the recent discovery that for a periodic configuration of scatterers the linear Boltzmann equation fails, and the random flight process that emerges in the Boltzmann-Grad limit is substantially more complicated. The key ingredient in the description of the limiting stochastic process is the renormalization dynamics on the space of lattices, a powerful technique that has recently been successfully applied also to other open problems in mathematical physics, including KAM theory and quantum chaos. This lecture is based on joint work with Andreas Strömbergsson, Uppsala.

  5. The classic: Bone morphogenetic protein.

    Science.gov (United States)

    Urist, Marshall R; Strates, Basil S

    2009-12-01

    This Classic Article is a reprint of the original work by Marshall R. Urist and Basil S. Strates, Bone Morphogenetic Protein. An accompanying biographical sketch of Marshall R. Urist, MD is available at DOI 10.1007/s11999-009-1067-4; a second Classic Article is available at DOI 10.1007/s11999-009-1069-2; and a third Classic Article is available at DOI 10.1007/s11999-009-1070-9. The Classic Article is copyright 1971 by Sage Publications Inc. Journals and is reprinted with permission from Urist MR, Strates BS. Bone morphogenetic protein. J Dent Res. 1971;50:1392-1406.

  6. Revisiting a classic: the Parker-Moffatt problem

    CERN Document Server

    Pezzi, O; Servidio, S; Valentini, F; Vasconez, C L; Yang, Y; Malara, F; Matthaeus, W H; Veltri, P

    2016-01-01

    The interaction of two colliding Alfv\\'en wave packets is here described by means of magnetohydrodynamics (MHD) and hybrid kinetic numerical simulations. The MHD evolution revisits the theoretical insights described by Moffatt, Parker, Kraichnan, Chandrasekhar and Els\\"asser in which the oppositely propagating large amplitude wave packets interact for a finite time, initiating turbulence. However, the extension to include compressive and kinetic effects, while maintaining the gross characteristics of the simpler classic formulation, also reveals intriguing features which go beyond the pure MHD treatment.

  7. Revisiting a Classic: The Parker–Moffatt Problem

    Science.gov (United States)

    Pezzi, O.; Parashar, T. N.; Servidio, S.; Valentini, F.; Vásconez, C. L.; Yang, Y.; Malara, F.; Matthaeus, W. H.; Veltri, P.

    2017-01-01

    The interaction of two colliding Alfvén wave packets is described here by means of magnetohydrodynamics (MHD) and hybrid kinetic numerical simulations. The MHD evolution revisits the theoretical insights described by Moffatt, Parker, Kraichnan, Chandrasekhar, and Elsässer in which the oppositely propagating large-amplitude wave packets interact for a finite time, initiating turbulence. However, the extension to include compressive and kinetic effects, while maintaining the gross characteristics of the simpler classic formulation, also reveals intriguing features that go beyond the pure MHD treatment.

  8. Innovation: the classic traps.

    Science.gov (United States)

    Kanter, Rosabeth Moss

    2006-11-01

    Never a fad, but always in or out of fashion, innovation gets rediscovered as a growth enabler every half dozen years. Too often, though, grand declarations about innovation are followed by mediocre execution that produces anemic results, and innovation groups are quietly disbanded in cost-cutting drives. Each managerial generation embarks on the same enthusiastic quest for the next new thing. And each generation faces the same vexing challenges- most of which stem from the tensions between protecting existing revenue streams critical to current success and supporting new concepts that may be crucial to future success. In this article, Harvard Business School professor Rosabeth Moss Kanter reflects on the four major waves of innovation enthusiasm she's observed over the past 25 years. She describes the classic mistakes companies make in innovation strategy, process, structure, and skills assessment, illustrating her points with a plethora of real-world examples--including AT&T Worldnet, Timberland, and Ocean Spray. A typical strategic blunder is when managers set their hurdles too high or limit the scope of their innovation efforts. Quaker Oats, for instance, was so busy in the 1990s making minor tweaks to its product formulas that it missed larger opportunities in distribution. A common process mistake is when managers strangle innovation efforts with the same rigid planning, budgeting, and reviewing approaches they use in their existing businesses--thereby discouraging people from adapting as circumstances warrant. Companies must be careful how they structure fledgling entities alongside existing ones, Kanter says, to avoid a clash of cultures and agendas--which Arrow Electronics experienced in its attempts to create an online venture. Finally, companies commonly undervalue and underinvest in the human side of innovation--for instance, promoting individuals out of innovation teams long before their efforts can pay off. Kanter offers practical advice for avoiding

  9. Non-equilibrium thermodynamics and physical kinetics

    CERN Document Server

    Bikkin, Halid

    2014-01-01

    This graduate textbook covers contemporary directions of non-equilibrium statistical mechanics as well as classical methods of kinetics. With one of the main propositions being to avoid terms such as "obviously" and "it is easy to show", this treatise is an easy-to-read introduction into this traditional, yet vibrant field.

  10. Exploring the Altered Dynamics of Mammalian Central Carbon Metabolic Pathway in Cancer Cells: A Classical Control Theoretic Approach.

    Directory of Open Access Journals (Sweden)

    Debjyoti Paul

    Full Text Available In contrast with normal cells, most of the cancer cells depend on aerobic glycolysis for energy production in the form of adenosine triphosphate (ATP bypassing mitochondrial oxidative phosphorylation. Moreover, compared to normal cells, cancer cells exhibit higher consumption of glucose with higher production of lactate. Again, higher rate of glycolysis provides the necessary glycolytic intermediary precursors for DNA, protein and lipid synthesis to maintain high active proliferation of the tumor cells. In this scenario, classical control theory based approach may be useful to explore the altered dynamics of the cancer cells. Since the dynamics of the cancer cells is different from that of the normal cells, understanding their dynamics may lead to development of novel therapeutic strategies.We have developed a model based on the state space equations of classical control theory along with an order reduction technique to mimic the actual dynamic behavior of mammalian central carbon metabolic (CCM pathway in normal cells. Here, we have modified Michaelis Menten kinetic equation to incorporate feedback mechanism along with perturbations and cross talks associated with a metabolic pathway. Furthermore, we have perturbed the proposed model to reduce the mitochondrial oxidative phosphorylation. Thereafter, we have connected proportional-integral (PI controller(s with the model for tuning it to behave like the CCM pathway of a cancer cell. This methodology allows one to track the altered dynamics mediated by different enzymes.The proposed model successfully mimics all the probable dynamics of the CCM pathway in normal cells. Moreover, experimental results demonstrate that in cancer cells, a coordination among enzymes catalyzing pentose phosphate pathway and intermediate glycolytic enzymes along with switching of pyruvate kinase (M2 isoform plays an important role to maintain their altered dynamics.

  11. A New Perspective on Classical Ideal Gases

    Directory of Open Access Journals (Sweden)

    Fabrice Philippe

    2013-08-01

    Full Text Available The ideal-gas barometric and pressure laws are derived from the Democritian concept of independent corpuscles moving in vacuum, plus a principle of simplicity, namely that these laws are independent of the kinetic part of the Hamiltonian. A single corpuscle in contact with a heat bath in a cylinder and submitted to a constant force (weight is considered. The paper importantly supplements a previously published paper: First, the stability of ideal gases is established. Second, we show that when walls separate the cylinder into parts and are later removed, the entropy is unaffected. We obtain full agreement with Landsberg’s and others’ (1994 classical thermodynamic result for the entropy of a column of gas submitted to gravity.

  12. The equilibrium classical scatter spectrum of waves

    CERN Document Server

    Guruprasad, V

    2016-01-01

    Regardless of the unspecific notions of photons as light complexes, radiation bundles or wave packets, the radiation from a single state transition is at most a single continuous wave train that starts and ends with the transition. The radiation equilibrium spectrum must be the superposition sum of the spectra of such wave trains. A classical equipartition of wave trains cannot diverge since they would be finite in number, whereas standing wave modes are by definition infinite, which had doomed Rayleigh's theory, and concern only the total radiation. Wave trains are the microscopic entities of radiation interacting with matter, that correspond to molecules in kinetic theory. Their quantization came from matter transitions in Einstein's 1917 derivation of Planck's law. The spectral scatter of wave trains by Doppler shifts, which cause the wavelength displacements in Wien's law used for the frequency dependence in Einstein's derivation, is shown to yield the shape of the Planck spectrum. A Lorentz transform pro...

  13. Structural and kinetic insights reveal that the amino acid pair Gln-228/Asn-254 modulates the transfructosylating specificity of Schwanniomyces occidentalis β-fructofuranosidase, an enzyme that produces prebiotics.

    Science.gov (United States)

    Álvaro-Benito, Miguel; Sainz-Polo, M Angela; González-Pérez, David; González, Beatriz; Plou, Francisco J; Fernández-Lobato, María; Sanz-Aparicio, Julia

    2012-06-01

    Schwanniomyces occidentalis β-fructofuranosidase (Ffase) is a GH32 dimeric enzyme that releases fructose from the nonreducing end of various oligosaccharides and essential storage fructans such as inulin. It also catalyzes the transfer of a fructosyl unit to an acceptor producing 6-kestose and 1-kestose, prebiotics that stimulate the growth of bacteria beneficial for human health. We report here the crystal structure of inactivated Ffase complexed with fructosylnystose and inulin, which shows the intricate net of interactions keeping the substrate tightly bound at the active site. Up to five subsites were observed, the sugar unit located at subsite +3 being recognized by interaction with the β-sandwich domain of the adjacent subunit within the dimer. This explains the high activity observed against long substrates, giving the first experimental evidence of the direct role of a GH32 β-sandwich domain in substrate binding. Crucial residues were mutated and their hydrolase/transferase (H/T) activities were fully characterized, showing the involvement of the Gln-228/Asn-254 pair in modulating the H/T ratio and the type β(2-1)/β(2-6) linkage formation. We generated Ffase mutants with new transferase activity; among them, Q228V gives almost specifically 6-kestose, whereas N254T produces a broader spectrum product including also neokestose. A model for the mechanism of the Ffase transfructosylation reaction is proposed. The results contribute to an understanding of the molecular basis regulating specificity among GH-J clan members, which represent an interesting target for rational design of enzymes, showing redesigned activities to produce tailor-made fructooligosaccharides.

  14. Atypical cytochrome p450 kinetics: implications for drug discovery.

    Science.gov (United States)

    Tracy, Timothy S

    2006-01-01

    The Michaelis-Menten model is commonly used to estimate a drug's potential in vivo hepatic clearance based on in vitro data obtained during drug discovery and development. This paradigm assumes that the drug obeys 'typical' enzyme kinetics and thus can be described by this model. However, it is increasingly being recognised that a number of drugs metabolised not only by the cytochrome P450 enzymes but also by other enzymes and transporters can exhibit atypical kinetic profiles, and thus are not accurately modeled with the Michaelis-Menten model. Application of an incorrect model can then lead to mis-estimation of in vitro intrinsic clearance and thus affect the prediction of in vivo clearance. This review discusses several types of atypical kinetic profiles that may be observed, including examples of homotropic cooperativity (i.e. sigmoidal kinetics, biphasic kinetics and substrate inhibition kinetics) as well as heterotropic cooperativity (i.e. activation). Application of the incorrect kinetic model may profoundly affect estimations of intrinsic clearance. For example, incorrectly applying the Michaelis-Menten model to a kinetic profile exhibiting substrate inhibition kinetics will result in an underestimation of Km (Michaelis-Menten constant) and V(max) (maximal velocity), whereas application of the Michaelis-Menten model to sigmoidal kinetic data typically results in an overestimation of Km and V(max) at the lower substrate concentrations that are typically therapeutically relevant. One must also be careful of potential artefactual causes of atypical kinetic profiles, such as enzyme activation by solvents, buffer dependent kinetic profiles, or altered kinetic parameter estimates due to nonspecific binding of the substrate to proteins. Despite a plethora of data on the effects of atypical kinetic profiles in vitro, only modest effects have been noted in vivo (with the exception of substrate dependent inhibition). Thus, the clinical relevance of these phenomena

  15. Operator Formulation of Classical Mechanics.

    Science.gov (United States)

    Cohn, Jack

    1980-01-01

    Discusses the construction of an operator formulation of classical mechanics which is directly concerned with wave packets in configuration space and is more similar to that of convential quantum theory than other extant operator formulations of classical mechanics. (Author/HM)

  16. Dynamical Symmetries in Classical Mechanics

    Science.gov (United States)

    Boozer, A. D.

    2012-01-01

    We show how symmetries of a classical dynamical system can be described in terms of operators that act on the state space for the system. We illustrate our results by considering a number of possible symmetries that a classical dynamical system might have, and for each symmetry we give examples of dynamical systems that do and do not possess that…

  17. Teaching and Demonstrating Classical Conditioning.

    Science.gov (United States)

    Sparrow, John; Fernald, Peter

    1989-01-01

    Discusses classroom demonstrations of classical conditioning and notes tendencies to misrepresent Pavlov's procedures. Describes the design and construction of the conditioner that is used for demonstrating classical conditioning. Relates how students experience conditioning, generalization, extinction, discrimination, and spontaneous recovery.…

  18. Classic writings on instructional technology

    NARCIS (Netherlands)

    Ely, Donald P.; Plomp, Tjeerd

    1996-01-01

    This paper describes the selection process of 17 articles for inclusion in the book, "Classic Writings on Instructional Technology." The book brings together original "classic" educational technology articles into one volume to document the history of the field through its literature. It is also an

  19. COMPETITION: CLASSICAL VERSUS NEOCLASSICAL VIEW

    OpenAIRE

    Mihaela Cornelia Sandu

    2013-01-01

    Competition is an important element from economical theory. Over time it has experienced several definitions and classifications much of them being contradictory. In this paper I will make a parallel between classical and neoclassical point of view according to competition. Keywords. Competition; neoclassical theory; classical theory; monopolistic; perfect competition.

  20. Classic African American Children's Literature

    Science.gov (United States)

    McNair, Jonda C.

    2010-01-01

    The purpose of this article is to assert that there are classic African American children's books and to identify a sampling of them. The author presents multiple definitions of the term classic based on the responses of children's literature experts and relevant scholarship. Next, the manner in which data were collected and analyzed in regard to…

  1. Classical Music Fan Chen Li

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The heyday of Beijing’s classical music beganin 1993, when top-quality sound equipment andrecords were imported. Also in that year, BeijingMusic Radio presented a classical music programtitled "Fan’s Club" and founded the "Music and

  2. Buprenorphine kinetics.

    Science.gov (United States)

    Bullingham, R E; McQuay, H J; Moore, A; Bennett, M R

    1980-11-01

    Buprenorphine kinetics was determined in surgical patients using radioimmunoassay. Buprenorphine was measured in the plasma of 24 patients who had received 0.3 mg buprenorphine intraoperatively. After 3 hr 10 of these patients then received a further 0.3 mg buprenorphine intravenously for postoperative pain relief, and 11 patients were given 0.3 mg intramuscularly; again, plasma levels were measured for 3 hr. The data fitted closely to a triexponential decay curve. There was a very fast initial phase, with a half-life (t1/2) of 2 min. The terminal t1/2 was slow, approximately 3 hr. Comparison of the kinetics of the same patient, awake and anesthetized, showed that the clearance was significantly lower in the anesthetized state. A notable feature of the drug given intramuscularly is rapid systemic availability, so that peaks are obtained in 2 to 5 min, and in 10 min the resulting levels are the same as for the intravenous and intramuscular routes.

  3. Classical approach in quantum physics

    CERN Document Server

    Solov'ev, Evgeni A

    2010-01-01

    The application of a classical approach to various quantum problems - the secular perturbation approach to quantization of a hydrogen atom in external fields and a helium atom, the adiabatic switching method for calculation of a semiclassical spectrum of hydrogen atom in crossed electric and magnetic fields, a spontaneous decay of excited states of a hydrogen atom, Gutzwiller's approach to Stark problem, long-lived excited states of a helium atom recently discovered with the help of Poincar$\\acute{\\mathrm{e}}$ section, inelastic transitions in slow and fast electron-atom and ion-atom collisions - is reviewed. Further, a classical representation in quantum theory is discussed. In this representation the quantum states are treating as an ensemble of classical states. This approach opens the way to an accurate description of the initial and final states in classical trajectory Monte Carlo (CTMC) method and a purely classical explanation of tunneling phenomenon. The general aspects of the structure of the semicla...

  4. Classical dynamics a modern perspective

    CERN Document Server

    Sudarshan, Ennackal Chandy George

    2016-01-01

    Classical dynamics is traditionally treated as an early stage in the development of physics, a stage that has long been superseded by more ambitious theories. Here, in this book, classical dynamics is treated as a subject on its own as well as a research frontier. Incorporating insights gained over the past several decades, the essential principles of classical dynamics are presented, while demonstrating that a number of key results originally considered only in the context of quantum theory and particle physics, have their foundations in classical dynamics.Graduate students in physics and practicing physicists will welcome the present approach to classical dynamics that encompasses systems of particles, free and interacting fields, and coupled systems. Lie groups and Lie algebras are incorporated at a basic level and are used in describing space-time symmetry groups. There is an extensive discussion on constrained systems, Dirac brackets and their geometrical interpretation. The Lie-algebraic description of ...

  5. Enzyme detection by microfluidics

    DEFF Research Database (Denmark)

    2013-01-01

    Microfluidic-implemented methods of detecting an enzyme, in particular a DNA-modifying enzyme, are provided, as well as methods for detecting a cell, or a microorganism expressing said enzyme. The enzyme is detected by providing a nucleic acid substrate, which is specifically targeted...... by that enzyme...

  6. Kinetic buffers.

    Science.gov (United States)

    Alibrandi, Giuseppe; Fabbrizzi, Luigi; Licchelli, Maurizio; Puglisi, Antonio

    2015-01-12

    This paper proposes a new type of molecular device that is able to act as an inverse proton sponge to slowly decrease the pH inside a reaction vessel. This makes the automatic monitoring of the concentration of pH-sensitive systems possible. The device is a composite formed of an alkyl chloride, which kinetically produces acidity, and a buffer that thermodynamically modulates the variation in pH value. Profiles of pH versus time (pH-t plots) have been generated under various experimental conditions by computer simulation, and the device has been tested by carrying out automatic spectrophotometric titrations, without using an autoburette. To underline the wide variety of possible applications, this new system has been used to realize and monitor HCl uptake by a di-copper(II) bistren complex in a single run, in a completely automatic experiment.

  7. eduction for Michaelis-Menten-Henri kinetics in the presence of diffusion

    Directory of Open Access Journals (Sweden)

    Leonid V. Kalachev

    2007-05-01

    Full Text Available The Michaelis-Menten-Henri (MMH mechanism is one of the paradigm reaction mechanisms in biology and chemistry. In its simplest form, it involves a substrate that reacts (reversibly with an enzyme, forming a complex which is transformed (irreversibly into a product and the enzyme. Given these basic kinetics, a dimension reduction has traditionally been achieved in two steps, by using conservation relations to reduce the number of species and by exploiting the inherent fast-slow structure of the resulting equations. In the present article, we investigate how the dynamics change if the species are additionally allowed to diffuse. We study the two extreme regimes of large diffusivities and of small diffusivities, as well as an intermediate regime in which the time scale of diffusion is comparable to that of the fast reaction kinetics. We show that reduction is possible in each of these regimes, with the nature of the reduction being regime dependent. Our analysis relies on the classical method of matched asymptotic expansions to derive approximations for the solutions that are uniformly valid in space and time.

  8. Plasmon mass scale in classical nonequilibrium gauge theory

    CERN Document Server

    Lappi, Tuomas

    2016-01-01

    Classical lattice Yang-Mills calculations provide a good way to understand different nonequilibrium phenomena in nonperturbatively overoccupied systems. Above the Debye scale the classical theory can be matched smoothly to kinetic theory. The aim of this work is to study the limits of this quasiparticle picture by determining the plasmon mass in classical real time Yang-Mills theory on a lattice in 3 spatial dimensions. We compare three methods to determine the plasmon mass: a hard thermal loop expression in terms of the particle distribution, an effective dispersion relation constructed from fields and their time derivatives, and by measuring oscillations between electric and magnetic field modes after artificially introducing a homogeneous color electric field. We find that a version of the dispersion relation that uses electric fields and their time derivatives agrees with the other methods within 50%.

  9. Classical Knowledge for Quantum Security

    CERN Document Server

    D'Hondt, Ellie

    2008-01-01

    We propose a decision procedure for analysing security of quantum cryptographic protocols, combining a classical algebraic rewrite system for knowledge with an operational semantics for quantum distributed computing. As a test case, we use our procedure to reason about security properties of a recently developed quantum secret sharing protocol that uses graph states. We analyze three different scenarios based on the safety assumptions of the classical and quantum channels and discover the path of an attack in the presence of an adversary. The epistemic analysis that leads to this and similar types of attacks is purely based on our classical notion of knowledge.

  10. Quantum localization of Classical Mechanics

    CERN Document Server

    Batalin, Igor A

    2016-01-01

    Quantum localization of classical mechanics within the BRST-BFV and BV (or field-antifield) quantization methods are studied. It is shown that a special choice of gauge fixing functions (or BRST-BFV charge) together with the unitary limit leads to Hamiltonian localization in the path integral of the BRST-BFV formalism. In turn, we find that a special choice of gauge fixing functions being proportional to extremals of an initial non-degenerate classical action together with a very special solution of the classical master equation result in Lagrangian localization in the partition function of the BV formalism.

  11. Quantum localization of classical mechanics

    Science.gov (United States)

    Batalin, Igor A.; Lavrov, Peter M.

    2016-07-01

    Quantum localization of classical mechanics within the BRST-BFV and BV (or field-antifield) quantization methods are studied. It is shown that a special choice of gauge fixing functions (or BRST-BFV charge) together with the unitary limit leads to Hamiltonian localization in the path integral of the BRST-BFV formalism. In turn, we find that a special choice of gauge fixing functions being proportional to extremals of an initial non-degenerate classical action together with a very special solution of the classical master equation result in Lagrangian localization in the partition function of the BV formalism.

  12. The Wigner representation of classical mechanics, quantization and classical limit

    Energy Technology Data Exchange (ETDEWEB)

    Bolivar, A.O. [Centro Brasileiro de Pesquisas Fisicas (CBPF), Rio de Janeiro, RJ (Brazil)

    2001-08-01

    Starting from the Liouvillian formulation of classical physics it is possible by means of a Fourier transform to introduce the Wigner representation and to derive an operator structure to classical mechanisms. The importance of this new representation lies on the fact that it turns out to be suitable route to establish a general method of quantization directly from the equations of motion without alluding to the existence of Hamiltonian and Lagrangian functions. Following this approach we quantize only the motion of a Browian particle with non-linear friction in the Markovian approximation - the thermal bath may be quantum or classical -, thus when the bath is classically described we obtain a master equation which reduces to Caldeira-Legget equation for the linear friction case, and when the reservoir is quantum we get an equation reducing to the one found by Caldeira et al. By neglecting the environmental influence we show that the system can be approximately described by equations of motion in terms of wave function, such as the Schrodinger-Langevin equation and equations of the Caldirola-Kanai type. Finally to make the present study self-consistent we evaluate the classical limit of these dynamical equations employing a new classical limiting method h/2{pi} {yields} 0. (author)

  13. Elevated Liver Enzymes

    Science.gov (United States)

    Symptoms Elevated liver enzymes By Mayo Clinic Staff Elevated liver enzymes may indicate inflammation or damage to cells in the liver. Inflamed or ... than normal amounts of certain chemicals, including liver enzymes, into the bloodstream, which can result in elevated ...

  14. New perspectives on classical electromagnetism

    OpenAIRE

    Cote, Paul J.

    2009-01-01

    The fallacies associated with the gauge concept in electromagnetism are illustrated. A clearer and more valid formulation of the basics of classical electromagnetism is provided by recognizing existing physical constraints as well as the physical reality of the vector potential.

  15. Soundscape of classical Chinese garden

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    With deep humanized connotation,the classical Chinese garden uses human intuitive sensation and personal poetic observation to express natural sound phenomena.It differs from the rational modern soundscape in western countries.

  16. Invariants from classical field theory

    CERN Document Server

    Diaz, Rafael

    2007-01-01

    We introduce a method that generates invariant functions from classical field theories depending on external parameters. We apply our method to several field theories such as abelian BF, Chern-Simons and 2-dimensional Yang-Mills theory.

  17. From Classical to Quantum Transistor

    OpenAIRE

    Sanjeev Kumar

    2009-01-01

    In this article the classical transistor and the basic physics underlying the operation of single electron transistor are presented; a brief history of transistor and current technological issues are discussed.

  18. From Classical to Quantum Transistor

    Directory of Open Access Journals (Sweden)

    Sanjeev Kumar

    2009-05-01

    Full Text Available In this article the classical transistor and the basic physics underlying the operation of single electron transistor are presented; a brief history of transistor and current technological issues are discussed.

  19. Classical Mechanics and Symplectic Integration

    DEFF Research Database (Denmark)

    Nordkvist, Nikolaj; Hjorth, Poul G.

    2005-01-01

    Content: Classical mechanics: Calculus of variations, Lagrange’s equations, Symmetries and Noether’s theorem, Hamilton’s equations, cannonical transformations, integrable systems, pertubation theory. Symplectic integration: Numerical integrators, symplectic integrators, main theorem on symplectic...

  20. Study on Metabolism Enzyme Kinetics of a Natural Novel Tyrosinase Inhibitor UP302 in Rat Liver Microsome by LC-MS/MS%LC-MS/MS研究新型酪氨酸酶抑制剂UP302在大鼠肝微粒体中代谢的酶动力学

    Institute of Scientific and Technical Information of China (English)

    朱凌; 张双庆; 闻镍; 于敏; 李佐刚

    2011-01-01

    OBJECTIVE To establish and validate a LC-MS/MS method for the determination of UP302 in rat liver microsome, and study the enzyme kinetics of UP3O2 metabolism in raHiver mierosome. METHODS The LC-MS/MS method with mass transitions of m/z 301. 1->-135. 2 for UP302 and 252. 9->132. 0 for internal standard daidzein was employed to determine UP302 incubated in rat liver microsome at negative electrospray ionization mode. Different incubation conditions, such as incubation time, substrate concentration and microsome concentration, were optimized. RESULTS The linear range of the calibration curve was 0. 1 - 20 (j.mol mL~' with good linear relationship. The intra- and inter-day accuracy was 86.42% -112. 59% , and RSDs were less than 9. 09%. The enzyme kinetic parameters were as follows:maximum reaction speed I^, of 196. 08 (junol mL"' Min"1 G"' , Km of 14. 98 pjnol mL"1, and CLiM of 13. 09 mL"1 Min"1 G~\\ CONCLUSION The established LC-MS/MS method was fast, simple, specific, sensitive and successfully applied to investigate the enzyme kinetics of UP302 metabolism in rat liver microsome.%目的 建立并验证测定大鼠肝微粒体孵育液中UP302含量的高效液相色谱-串联质谱法,研究UP302在大鼠肝微粒体中代谢的酶动力学.方法 经大鼠肝微粒体孵育的UP302样品,经色谱柱分离,电喷雾离子化负离子检测,扫描方式为选择反应监测,用于定量分析的离子反应分别为m/z 301.1→135.2(UP302)和m/z252.9→132.O(内标大豆苷元).考察不同孵育时间、底物浓度和微粒体浓度对UP302代谢的影响,确定最佳反应条件.结果 标准曲线线性范围为0.1~20 μmol·L-1,线性关系良好,日内日间准确度在86.42%~ 112.59%,日内日间精密度均小于9.09%,回收率在100.50% ~ 109.91%之间.确定了酶动力学参数,最大反应速度Vmax为196.08 μmol·L-1·min-1·g-1,米氏常数km为14.98μmol·L-1,内在代谢清除率CLint为13.09 mL-1·min-1·g-1.结论 该检测方法

  1. Electrostatics interactions in classical simulations.

    Science.gov (United States)

    Cisneros, G Andrés; Babin, Volodymyr; Sagui, Celeste

    2013-01-01

    Electrostatic interactions are crucial for both the accuracy and performance of atomistic biomolecular simulations. In this chapter we review well-established methods and current developments aiming at efficiency and accuracy. Specifically, we review the classical Ewald summations, particle-particle particle-method particle-method Ewald algorithms, multigrid, fast multipole, and local methods. We also highlight some recent developments targeting more accurate, yet classical, representation of the molecular charge distribution.

  2. Classical Transitions for Flux Vacua

    CERN Document Server

    Deskins, J Tate; Yang, I-Sheng

    2012-01-01

    We present the simplest model for classical transitions in flux vacua. A complex field with a spontaneously broken U(1) symmetry is embedded in $M_2\\times S_1$. We numerically construct different winding number vacua, the vortices interpolating between them, and simulate the collisions of these vortices. We show that classical transitions are generic at large boosts, independent of whether or not vortices miss each other in the compact $S_1$.

  3. New Perspective on Classical Electromagnetism

    Science.gov (United States)

    2013-04-01

    R. Feynman , R. Leighton, and M. Sands, The Feynman Lectures in Physics vol II (Addison-Wesley, Reading, MA, 1964). 6. W.K.H. Panofsky and M...of the basics of classical electromagnetism is provided by recognizing a previously overlooked law of induction as well as the physical reality of the...classical electromagnetism is provided by recognizing a previously overlooked law of induction as well as the physical reality of the vector potential

  4. Classical theory of radiating strings

    Science.gov (United States)

    Copeland, Edmund J.; Haws, D.; Hindmarsh, M.

    1990-01-01

    The divergent part of the self force of a radiating string coupled to gravity, an antisymmetric tensor and a dilaton in four dimensions are calculated to first order in classical perturbation theory. While this divergence can be absorbed into a renormalization of the string tension, demanding that both it and the divergence in the energy momentum tensor vanish forces the string to have the couplings of compactified N = 1 D = 10 supergravity. In effect, supersymmetry cures the classical infinities.

  5. Examinations of the Chemical Step in Enzyme Catalysis.

    Science.gov (United States)

    Singh, P; Islam, Z; Kohen, A

    2016-01-01

    Advances in computational and experimental methods in enzymology have aided comprehension of enzyme-catalyzed chemical reactions. The main difficulty in comparing computational findings to rate measurements is that the first examines a single energy barrier, while the second frequently reflects a combination of many microscopic barriers. We present here intrinsic kinetic isotope effects and their temperature dependence as a useful experimental probe of a single chemical step in a complex kinetic cascade. Computational predictions are tested by this method for two model enzymes: dihydrofolate reductase and thymidylate synthase. The description highlights the significance of collaboration between experimentalists and theoreticians to develop a better understanding of enzyme-catalyzed chemical conversions.

  6. Orphan enzymes in ether lipid metabolism.

    Science.gov (United States)

    Watschinger, Katrin; Werner, Ernst R

    2013-01-01

    Ether lipids are an emerging class of lipids which have so far not been investigated and understood in every detail. They have important roles as membrane components of e.g. lens, brain and testis, and as mediators such as platelet-activating factor. The metabolic enzymes for biosynthesis and degradation have been investigated to some extent. As most involved enzymes are integral membrane proteins they are tricky to handle in biochemical protocols. The sequence of some ether lipid metabolising enzymes has only recently been reported and other sequences still remain obscure. Defined enzymes without assigned sequence are known as orphan enzymes. One of these enzymes with uncharacterised sequence is plasmanylethanolamine desaturase, a key enzyme for the biosynthesis of one of the most abundant phospholipids in our body, the plasmalogens. This review aims to briefly summarise known functions of ether lipids, give an overview on their metabolism including the most prominent members, platelet-activating factor and the plasmalogens. A special focus is set on the description of orphan enzymes in ether lipid metabolism and on the successful strategies how four previous orphans have recently been assigned a sequence. Only one of these four was characterised by classical protein purification and sequencing, whereas the other three required alternative strategies such as bioinformatic candidate gene selection and recombinant expression or development of an inhibitor and multidimensional metabolic profiling.

  7. Bisphenol A degradation in water by ligninolytic enzymes.

    Science.gov (United States)

    Gassara, Fatma; Brar, Satinder K; Verma, M; Tyagi, R D

    2013-08-01

    Many endocrine disruptor compounds, such as bisphenol A (BPA) are used today and released into the environment at low doses but they are barely degraded in wastewater treatment plants. One of the potential alternatives to effectively degrade endocrine disruptor compounds is based on the use of the oxidative action of extracellular fungal enzymes. The aim of this work is to study the ability of free and encapsulated enzymes (manganese peroxidase, lignin peroxidase and laccase) to degrade BPA. Higher degradation of BPA (90%) by ligninolytic enzymes encapsulated on polyacrylamide hydrogel and pectin after 8h was obtained. The degradation of BPA while using the free enzyme (26%) was lower than the value obtained with encapsulated enzymes. The presence of pectin in the formulation significantly (p>0.05) enhanced the activity of enzymes. Kinetics of BPA degradation showed an increase in Vm, while Km remained constant when enzymes were encapsulated. Hence, encapsulation protected the enzymes from non-competitive inhibition.

  8. The Kinetics of Carrier Transport Inhibition

    DEFF Research Database (Denmark)

    Rosenberg, T.; Wilbrandt, Robert Walter

    1962-01-01

    The kinetical treatment of enzymatic carrier transports as given in previous communications has been extended to conditions of inhibition. Various possible types of inhibitors have been considered differing in the site of attack (enzyme or carrier), in the mode of action (competing...... with the substrate for the enzyme or the carrier or for both, competing with the carrier for the enzyme, or non-competitive) and in the ability of penetrating the membrane. Experiments are reported on the inhibition of glucose and fructose transport across the human red cell membrane by phlorizine, phloretine...... the first order asymmetry severalfold (“second order asymmetry”). It was shown that a substrate competitive mode of action involving competition both for the enzyme and for the enzyme-bound carrier will result in a behaviour resembling the observed “second order asymmetry”. It is felt, therefore...

  9. Enzymes: The possibility of production and applications

    Directory of Open Access Journals (Sweden)

    Petronijević Živomir B.

    2003-01-01

    Full Text Available Enzymes are biological catalysts with increasing application in the food pharmaceutical, cosmetic, textile and chemical industry. They are also important as reagents in chemical analysis, leather fabrications and as targets for the design of new drugs. Keeping in mind the growing need to replace classical chemical processes by alternative ones, because of ever growing environmental pollution, it is important that enzyme and other biotechnological processes are economical. Therefore, price decrease and stability and enzyme preparation efficiency increase are required more and more. This paper presents a short review of methods for yield increase and the improvement of the quality of enzyme products as commercial products, as well as a review of the possibilities of their application.

  10. Hermeneutic reading of classic texts.

    Science.gov (United States)

    Koskinen, Camilla A-L; Lindström, Unni Å

    2013-09-01

    The purpose of this article is to broaden the understandinfg of the hermeneutic reading of classic texts. The aim is to show how the choice of a specific scientific tradition in conjunction with a methodological approach creates the foundation that clarifies the actual realization of the reading. This hermeneutic reading of classic texts is inspired by Gadamer's notion that it is the researcher's own research tradition and a clearly formulated theoretical fundamental order that shape the researcher's attitude towards texts and create the starting point that guides all reading, uncovering and interpretation. The researcher's ethical position originates in a will to openness towards what is different in the text and which constantly sets the researcher's preunderstanding and research tradition in movement. It is the researcher's attitude towards the text that allows the text to address, touch and arouse wonder. Through a flexible, lingering and repeated reading of classic texts, what is different emerges with a timeless value. The reading of classic texts is an act that may rediscover and create understanding for essential dimensions and of human beings' reality on a deeper level. The hermeneutic reading of classic texts thus brings to light constantly new possibilities of uncovering for a new envisioning and interpretation for a new understanding of the essential concepts and phenomena within caring science.

  11. Does classical liberalism imply democracy?

    Directory of Open Access Journals (Sweden)

    David Ellerman

    2015-12-01

    Full Text Available There is a fault line running through classical liberalism as to whether or not democratic self-governance is a necessary part of a liberal social order. The democratic and non-democratic strains of classical liberalism are both present today—particularly in the United States. Many contemporary libertarians and neo-Austrian economists represent the non-democratic strain in their promotion of non-democratic sovereign city-states (start-up cities or charter cities. We will take the late James M. Buchanan as a representative of the democratic strain of classical liberalism. Since the fundamental norm of classical liberalism is consent, we must start with the intellectual history of the voluntary slavery contract, the coverture marriage contract, and the voluntary non-democratic constitution (or pactum subjectionis. Next we recover the theory of inalienable rights that descends from the Reformation doctrine of the inalienability of conscience through the Enlightenment (e.g. Spinoza and Hutcheson in the abolitionist and democratic movements. Consent-based governments divide into those based on the subjects’ alienation of power to a sovereign and those based on the citizens’ delegation of power to representatives. Inalienable rights theory rules out that alienation in favor of delegation, so the citizens remain the ultimate principals and the form of government is democratic. Thus the argument concludes in agreement with Buchanan that the classical liberal endorsement of sovereign individuals acting in the marketplace generalizes to the joint action of individuals as the principals in their own organizations.

  12. No Return to Classical Reality

    CERN Document Server

    Jennings, David

    2015-01-01

    At a fundamental level, the classical picture of the world is dead, and has been dead now for almost a century. Pinning down exactly which quantum phenomena are responsible for this has proved to be a tricky and controversial question, but a lot of progress has been made in the past few decades. We now have a range of precise statements showing that whatever the ultimate laws of Nature are, they cannot be classical. In this article, we review results on the fundamental phenomena of quantum theory that cannot be understood in classical terms. We proceed by first granting quite a broad notion of classicality, describe a range of quantum phenomena (such as randomness, discreteness, the indistinguishability of states, measurement-uncertainty, measurement-disturbance, complementarity, noncommutativity, interference, the no-cloning theorem, and the collapse of the wave-packet) that do fall under its liberal scope, and then finally describe some aspects of quantum physics that can never admit a classical understandi...

  13. Classical approach in atomic physics

    Science.gov (United States)

    Solov'ev, E. A.

    2011-12-01

    The application of a classical approach to various quantum problems - the secular perturbation approach to quantization of a hydrogen atom in external fields and a helium atom, the adiabatic switching method for calculation of a semiclassical spectrum of a hydrogen atom in crossed electric and magnetic fields, a spontaneous decay of excited states of a hydrogen atom, Gutzwiller's approach to Stark problem, long-lived excited states of a helium atom discovered with the help of Poincaré section, inelastic transitions in slow and fast electron-atom and ion-atom collisions - is reviewed. Further, a classical representation in quantum theory is discussed. In this representation the quantum states are treated as an ensemble of classical states. This approach opens the way to an accurate description of the initial and final states in classical trajectory Monte Carlo (CTMC) method and a purely classical explanation of tunneling phenomenon. The general aspects of the structure of the semiclassical series such as renormgroup symmetry, criterion of accuracy and so on are reviewed as well.

  14. Classical approach in atomic physics

    Energy Technology Data Exchange (ETDEWEB)

    Solov' ev, E.A. [Bogoliubov Laboratory of Theoretical Physics, Joint Institute for Nuclear Research, 141980 Dubna (Russian Federation)

    2011-12-15

    The application of a classical approach to various quantum problems - the secular perturbation approach to quantization of a hydrogen atom in external fields and a helium atom, the adiabatic switching method for calculation of a semiclassical spectrum of a hydrogen atom in crossed electric and magnetic fields, a spontaneous decay of excited states of a hydrogen atom, Gutzwiller's approach to Stark problem, long-lived excited states of a helium atom discovered with the help of Poincare section, inelastic transitions in slow and fast electron-atom and ion-atom collisions - is reviewed. Further, a classical representation in quantum theory is discussed. In this representation the quantum states are treated as an ensemble of classical states. This approach opens the way to an accurate description of the initial and final states in classical trajectory Monte Carlo (CTMC) method and a purely classical explanation of tunneling phenomenon. The general aspects of the structure of the semiclassical series such as renormalization group symmetry, criterion of accuracy and so on are reviewed as well. (author)

  15. Substrate-dependent kinetics in tyrosinase-based biosensing: amperometry vs. spectrophotometry

    NARCIS (Netherlands)

    Rassaei, L.; Cui, J.; Goluch, E.D.; Lemay, S.G.

    2012-01-01

    Despite the broad use of enzymes in electroanalytical biosensors, the influence of enzyme kinetics on the function of prototype sensors is often overlooked or neglected. In the present study, we employ amperometry as an alternative or complementary method to study the kinetics of tyrosinase, whose

  16. How thiamine diphosphate is activated in enzymes.

    Science.gov (United States)

    Kern, D; Kern, G; Neef, H; Tittmann, K; Killenberg-Jabs, M; Wikner, C; Schneider, G; Hübner, G

    1997-01-03

    The controversial question of how thiamine diphosphate, the biologically active form of vitamin B1, is activated in different enzymes has been addressed. Activation of the coenzyme was studied by measuring thermodynamics and kinetics of deprotonation at the carbon in the 2-position (C2) of thiamine diphosphate in the enzymes pyruvate decarboxylase and transketolase by use of nuclear magnetic resonance spectroscopy, proton/deuterium exchange, coenzyme analogs, and site-specific mutant enzymes. Interaction of a glutamate with the nitrogen in the 1'-position in the pyrimidine ring activated the 4'-amino group to act as an efficient proton acceptor for the C2 proton. The protein component accelerated the deprotonation of the C2 atom by several orders of magnitude, beyond the rate of the overall enzyme reaction. Therefore, the earlier proposed concerted mechanism or stabilization of a C2 carbanion can be excluded.

  17. Kinetic ELISA in Microfluidic Channels

    Directory of Open Access Journals (Sweden)

    Debashis Dutta

    2011-06-01

    Full Text Available In this article, we describe the kinetic ELISA of Blue Tongue and Epizootic Hemorrhagic Disease viral antibodies in microfluidic channels by monitoring the rate of generation of the enzyme reaction product under static conditions. It has been shown that this format of the immunoassay allows very reliable quantitation of the target species using inexpensive glass microchips and a standard epifluorescence microscope system coupled to a CCD camera. For the viral antibodies assayed here, the limit of detection (LOD for the analyte concentration in our microchips was established to be 3–5 times lower than that obtained on commercial microwell plates using a fiftieth of the sample volume and less than a third of the incubation time. Our analyses further show that when compared to the end-point ELISA format, the kinetic mode of this assay yields an improvement in the LOD by over an order of magnitude in microfluidic devices. This benefit is primarily realized as the observed variation in the background fluorescence (signal at the start of the enzyme reaction period was significantly larger than that in the rate of signal generation upon repeating these assays in different microchannels/microchips. Because the kinetic ELISA results depend only on the latter quantity, the noise level in them was substantially lower compared to that in its end-point counterpart in which the absolute fluorescence measurements are of greater significance. While a similar benefit was also recorded through implementation of kinetic ELISAs on the microwell platform, the improvement in LOD registered in that system was not as significant as was observed in the case of microfluidic assays.

  18. Population in the classic economics

    Directory of Open Access Journals (Sweden)

    Adnan Doğruyol

    2013-02-01

    Full Text Available Growth subject in economics is an important factor of development. Classic economics ecole indicates the population as main variable which tender of growth. On the other hand T. R. Malthus is known as economist who regards population as a problem and brings up it among the classical economists. However, Adam Smith is an intellectual who discussed population problem earlier on the classic economics theory. According to Adam Smith one of the main factors that realise the growth is labour. In addition to population made it established. The aim of this study is analyzing the mental relationship between Malthus whose name has been identified with relation between population-growth and Smith who discussed this subject first time but put it off on process of theorisation.

  19. Optimum Onager: The Classical Mechanics of a Classical Siege Engine

    Science.gov (United States)

    Denny, Mark

    2009-01-01

    The onager is a throwing weapon of classical antiquity, familiar to both the ancient Greeks and Romans. Here we analyze the dynamics of onager operation and derive the optimum angle for launching a projectile to its maximum range. There is plenty of scope for further considerations about increasing onager range, and so by thinking about how this…

  20. Overview of Classical Swine Fever (Hog Cholera, Classical Swine fever)

    Science.gov (United States)

    Classical swine fever is a contagious often fatal disease of pigs clinically characterized by high body temperature, lethargy, yellowish diarrhea, vomits and purple skin discoloration of ears, lower abdomen and legs. It was first described in the early 19th century in the USA. Later, a condition i...

  1. Classical planning and causal implicatures

    DEFF Research Database (Denmark)

    Blackburn, Patrick Rowan; Benotti, Luciana

    In this paper we motivate and describe a dialogue manager (called Frolog) which uses classical planning to infer causal implicatures. A causal implicature is a type of Gricean relation implicature, a highly context dependent form of inference. As we shall see, causal implicatures are important...... to generate clarification requests"; as a result we can model task-oriented dialogue as an interactive process locally structured by negotiation of the underlying task. We give several examples of Frolog-human dialog, discuss the limitations imposed by the classical planning paradigm, and indicate...

  2. Comparing classical and quantum equilibration

    CERN Document Server

    Malabarba, Artur S L; Short, Anthony J

    2016-01-01

    By using a physically-relevant and theory independent definition of measurement-based equilibration, we show quantitatively that equilibration is easier for quantum systems than for classical systems, in the situation where the initial state of the system is completely known (pure state). This shows that quantum equilibration is a fundamental, nigh unavoidable, aspect of physical systems, while classical equilibration relies on experimental ignorance. When the state is not completely known, a mixed state, this framework also shows quantum equilibration requires weaker conditions.

  3. Principal bundles the classical case

    CERN Document Server

    Sontz, Stephen Bruce

    2015-01-01

    This introductory graduate level text provides a relatively quick path to a special topic in classical differential geometry: principal bundles.  While the topic of principal bundles in differential geometry has become classic, even standard, material in the modern graduate mathematics curriculum, the unique approach taken in this text presents the material in a way that is intuitive for both students of mathematics and of physics. The goal of this book is to present important, modern geometric ideas in a form readily accessible to students and researchers in both the physics and mathematics communities, providing each with an understanding and appreciation of the language and ideas of the other.

  4. Quantum teleportation without classical channel

    Science.gov (United States)

    Al Amri, M.; Li, Zheng-Hong; Zubairy, M. Suhail

    2016-11-01

    For the first time, we show how quantum teleportation can be achieved without the assistance of classical channels. Our protocol does not need any pre-established entangled photon pairs beforehand. Just by utilizing quantum Zeno effect and couterfactual communication idea, we can achieve two goals; entangling a photon and an atom and also disentangling them by non-local interaction. Information is completely transferred from atom to photon with controllable disentanglement processes. More importantly, there is no need to confirm teleportation results via classical channels.

  5. Teaching Classical Mechanics Using Smartphones

    Science.gov (United States)

    Chevrier, Joel; Madani, Laya; Ledenmat, Simon; Bsiesy, Ahmad

    2013-01-01

    A number of articles published in this column have dealt with topics in classical mechanics. This note describes some additional examples employing a smartphone and the new software iMecaProf. Steve Jobs presented the iPhone as "perfect for gaming." Thanks to its microsensors connected in real time to the numerical world, physics…

  6. Supersymmetric classical mechanics: free case

    Energy Technology Data Exchange (ETDEWEB)

    Rodrigues, R. de Lima [Centro Brasileiro de Pesquisas Fisicas (CBPF), Rio de Janeiro, RJ (Brazil)]|[Paraiba Univ., Cajazeiras, PB (Brazil). Dept. de Ciencias Exatas e da Natureza]. E-mail: rafael@cfp.ufpb.br; Almeida, W. Pires de [Paraiba Univ., Cajazeiras, PB (Brazil). Dept. de Ciencias Exatas e da Natureza; Fonseca Neto, I. [Paraiba Univ., Campina Grande, PB (Brazil). Dept. de Fisica

    2001-06-01

    We present a review work on Supersymmetric Classical Mechanics in the context of a Lagrangian formalism, with N = 1-supersymmetry. We show that the N = 1 supersymmetry does not allow the introduction of a potencial energy term depending on a single commuting supercoordinate, {phi}(t;{theta}). (author)

  7. Agglomeration Economies in Classical Music

    DEFF Research Database (Denmark)

    Borowiecki, Karol Jan

    2015-01-01

    This study investigates agglomeration effects for classical music production in a wide range of cities for a global sample of composers born between 1750 and 1899. Theory suggests a trade-off between agglomeration economies (peer effects) and diseconomies (peer crowding). I test this hypothesis...

  8. Classical and quantum Coulomb crystals

    CERN Document Server

    Bonitz, M; Baumgartner, H; Henning, C; Filinov, A; Block, D; Arp, O; Piel, A; Kading, S; Ivanov, Y; Melzer, A; Fehske, H; Filinov, V

    2008-01-01

    Strong correlation effects in classical and quantum plasmas are discussed. In particular, Coulomb (Wigner) crystallization phenomena are reviewed focusing on one-component non-neutral plasmas in traps and on macroscopic two-component neutral plasmas. The conditions for crystal formation in terms of critical values of the coupling parameters and the distance fluctuations and the phase diagram of Coulomb crystals are discussed.

  9. Classical Syllogisms in Logic Teaching

    DEFF Research Database (Denmark)

    Øhrstrøm, Peter; Sandborg-Petersen, Ulrik; Thorvaldsen, Steinar

    2013-01-01

    This paper focuses on the challenges of introducing classical syllogisms in university courses in elementary logic and human reasoning. Using a program written in Prolog+CG, some empirical studies have been carried out involving three groups of students in Denmark; one group of philosophy student...

  10. On classical and quantum liftings

    CERN Document Server

    Accardi, L; Kossakowski, A; Matsuoka, T; Ohya, M

    2011-01-01

    We analyze the procedure of lifting in classical stochastic and quantum systems. It enables one to `lift' a state of a system into a state of `system+reservoir'. This procedure is important both in quantum information theory and the theory of open systems. We illustrate the general theory of liftings by a particular class related to so called circulant states.

  11. Relative Clauses in Classical Nahuatl

    Science.gov (United States)

    Langacker, Ronald W.

    1975-01-01

    Jane Rosenthal's paper on relative clauses in Classical Nahuatl is discussed, and it is argued that she misses an important generalization. An alternative analysis to a class of relative pronouns and new rules for the distribution of relative pronouns are proposed. (SC)

  12. On Classical and Quantum Cryptography

    CERN Document Server

    Volovich, I V; Volovich, Ya.I.

    2001-01-01

    Lectures on classical and quantum cryptography. Contents: Private key cryptosystems. Elements of number theory. Public key cryptography and RSA cryptosystem. Shannon`s entropy and mutual information. Entropic uncertainty relations. The no cloning theorem. The BB84 quantum cryptographic protocol. Security proofs. Bell`s theorem. The EPRBE quantum cryptographic protocol.

  13. Minimum signals in classical physics

    Institute of Scientific and Technical Information of China (English)

    邓文基; 许基桓; 刘平

    2003-01-01

    The bandwidth theorem for Fourier analysis on any time-dependent classical signal is shown using the operator approach to quantum mechanics. Following discussions about squeezed states in quantum optics, the problem of minimum signals presented by a single quantity and its squeezing is proposed. It is generally proved that all such minimum signals, squeezed or not, must be real Gaussian functions of time.

  14. Classical and molecular genetic mapping

    Science.gov (United States)

    A brief history of classical genetic mapping in soybean [Glycine max (L.) Merr.] is described. Detailed descriptions are given of the development of molecular genetic linkage maps based upon various types of DNA markers Like many plant and animal species, the first molecular map of soybean was bas...

  15. Classical Music as Enforced Utopia

    Science.gov (United States)

    Leech-Wilkinson, Daniel

    2016-01-01

    In classical music composition, whatever thematic or harmonic conflicts may be engineered along the way, everything always turns out for the best. Similar utopian thinking underlies performance: performers see their job as faithfully carrying out their master's (the composer's) wishes. The more perfectly they represent them, the happier the…

  16. Biocatalytic synthesis of pyruvate from DL-lactate with enzymes in Pseudomonas sp

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    A novel method of preparing pyruvate from DL-lactate catalyzed by enzymes from a bacterial strain of Pseudomonas sp. SM-6 was proposed. Catalytic processes of cell-free extract enzymes and immobilized enzymes were evaluated. The kinetic data were studied, too.

  17. No return to classical reality

    Science.gov (United States)

    Jennings, David; Leifer, Matthew

    2016-01-01

    At a fundamental level, the classical picture of the world is dead, and has been dead now for almost a century. Pinning down exactly which quantum phenomena are responsible for this has proved to be a tricky and controversial question, but a lot of progress has been made in the past few decades. We now have a range of precise statements showing that whatever the ultimate laws of nature are, they cannot be classical. In this article, we review results on the fundamental phenomena of quantum theory that cannot be understood in classical terms. We proceed by first granting quite a broad notion of classicality, describe a range of quantum phenomena (such as randomness, discreteness, the indistinguishability of states, measurement-uncertainty, measurement-disturbance, complementarity, non-commutativity, interference, the no-cloning theorem and the collapse of the wave-packet) that do fall under its liberal scope, and then finally describe some aspects of quantum physics that can never admit a classical understanding - the intrinsically quantum mechanical aspects of nature. The most famous of these is Bell's theorem, but we also review two more recent results in this area. Firstly, Hardy's theorem shows that even a finite-dimensional quantum system must contain an infinite amount of information, and secondly, the Pusey-Barrett-Rudolph theorem shows that the wave function must be an objective property of an individual quantum system. Besides being of foundational interest, results of this sort now find surprising practical applications in areas such as quantum information science and the simulation of quantum systems.

  18. Does classical mechanics always adequately describe "classical physical reality"

    CERN Document Server

    Shemi-zadeh, V E

    2002-01-01

    The article is dedicated to discussion of irreversibility and foundation of statistical mechanics "from the first principles". Taking into account infinitesimal and, as it seems, neglectful for classical mechanics fluctuations of the physical vacuum, makes a deterministic motion of unstable dynamic systems is broken ("spontaneous determinism breaking", "spontaneous stochastization"). Vacuum fluctuations play part of the trigger, starting the powerful mechanism of exponent instability. The motion of the dynamic systems becomes irreversible and stochastic. Classical mechanics turns out to be applicable only for a small class of stable dynamic systems with zero Kolmogorov-Sinay entropy $h=0$. For alternative "Stochastic mechanics" there are corresponding equations of motion and Master Equation, describing irreversible evolution of the initial distribution function to equilibrium state.

  19. Enzyme dynamics from NMR spectroscopy.

    Science.gov (United States)

    Palmer, Arthur G

    2015-02-17

    CONSPECTUS: Biological activities of enzymes, including regulation or coordination of mechanistic stages preceding or following the chemical step, may depend upon kinetic or equilibrium changes in protein conformations. Exchange of more open or flexible conformational states with more closed or constrained states can influence inhibition, allosteric regulation, substrate recognition, formation of the Michaelis complex, side reactions, and product release. NMR spectroscopy has long been applied to the study of conformational dynamic processes in enzymes because these phenomena can be characterized over multiple time scales with atomic site resolution. Laboratory-frame spin-relaxation measurements, sensitive to reorientational motions on picosecond-nanosecond time scales, and rotating-frame relaxation-dispersion measurements, sensitive to chemical exchange processes on microsecond-millisecond time scales, provide information on both conformational distributions and kinetics. This Account reviews NMR spin relaxation studies of the enzymes ribonuclease HI from mesophilic (Escherichia coli) and thermophilic (Thermus thermophilus) bacteria, E. coli AlkB, and Saccharomyces cerevisiae triosephosphate isomerase to illustrate the contributions of conformational flexibility and dynamics to diverse steps in enzyme mechanism. Spin relaxation measurements and molecular dynamics (MD) simulations of the bacterial ribonuclease H enzymes show that the handle region, one of three loop regions that interact with substrates, interconverts between two conformations. Comparison of these conformations with the structure of the complex between Homo sapiens ribonuclease H and a DNA:RNA substrate suggests that the more closed state is inhibitory to binding. The large population of the closed conformation in T. thermophilus ribonuclease H contributes to the increased Michaelis constant compared with the E. coli enzyme. NMR spin relaxation and fluorescence spectroscopy have characterized a

  20. Kinetic properties of cyanase.

    Science.gov (United States)

    Anderson, P M; Little, R M

    1986-04-01

    Cyanase is an inducible enzyme in Escherichia coli that catalyzes the hydrolysis of cyanate. Bicarbonate is required for activity, perhaps as a substrate, and the initial product of the reaction is carbamate, which spontaneously breaks down to ammonia and bicarbonate [Anderson, P. M. (1980) Biochemistry 19, 2882]. The purpose of this study was to characterize the kinetic properties of cyanase. Initial velocity studies showed that both cyanate and bicarbonate act as competitive substrate inhibitors. A number of monovalent anions act as inhibitors. Azide and acetate appear to act as competitive inhibitors with respect to cyanate and bicarbonate, respectively. Chloride, bromide, nitrate, nitrite, and formate also inhibit, apparently as the result of binding at either substrate site. Malonate and several other dicarboxylic dianions at very low concentrations display "slow-binding", reversible inhibition which can be prevented by saturating concentrations of either substrate. The results are consistent with a rapid equilibrium random mechanism in which bicarbonate acts as a substrate, bicarbonate and cyanate bind at adjacent anion-binding sites, and both substrates can bind at the other substrate anion binding site to give a dead-end complex.

  1. Classical trajectory Monte Carlo investigation for Lorentz ionization of H (1s)

    Institute of Scientific and Technical Information of China (English)

    He Bin; Wang Jian-Guo; Liu Chun-Lei

    2013-01-01

    Lorentz ionization of H(1s) is investigated by classical trajectory Monte Carlo (CTMC) simulation.The effect of the transverse magnetic field on the considered process is analyzed in terms of the time evolution of interactions in the system,total electron energy,and electron trajectories.A classical mechanism for the ionization is found,where the variation of the kinetic energy of the nuclei is found to be important in the process.Compared with the results of tunneling ionization,the classical mechanism becomes more and more important with the increase of the velocity of the H-atom or the strength of the magnetic field.

  2. Why Can We Copy Classical Information?

    Institute of Scientific and Technical Information of China (English)

    SHEN Yao; HAO Liang; LONG Gui-Lu

    2011-01-01

    It is pointed out that the noncloning theorem in quantum mechanics also holds for unknown state in linear classical physics. The apparent capability of copying of a classical state is essentially the capability of perfect measurement in classical physics. The difference in copying between quantum and classical physics is the difference in measurement between the two theories. A classical copying process is the combined action of measurement of an unknown state and the preparation of this state onto another system. Hence perfect measurability in classical physics enables the copying of a classical state.

  3. Descriptive and predictive assessment of enzyme activity and enzyme related processes in biorefinery using IR spectroscopy and chemometrics

    DEFF Research Database (Denmark)

    Baum, Andreas

    the understanding of the structural properties of the extracted pectin. Secondly, enzyme kinetics of biomass converting enzymes was examined in terms of measuring enzyme activity by spectral evolution profiling utilizing FTIR. Chemometric multiway methods were used to analyze the tensor datasets enabling the second......-order calibration advantage (reference Theory of Analytical chemistry). As PAPER 3 illustrates the method is universally applicable without the need of any external standards and was exemplified by performing quantitative enzyme activity determinations for glucose oxidase, pectin lyase and a cellolytic enzyme blend...... (Celluclast 1.5L). In PAPER 4, the concept is extended to quantify enzyme activity of two simultaneously acting enzymes, namely pectin lyase and pectin methyl esterase. By doing so the multiway methods PARAFAC, TUCKER3 and NPLS were compared and evaluated towards accuracy and precision....

  4. A method for analysis and design of metabolism using metabolomics data and kinetic models: Application on lipidomics using a novel kinetic model of sphingolipid metabolism.

    Science.gov (United States)

    Savoglidis, Georgios; da Silveira Dos Santos, Aline Xavier; Riezman, Isabelle; Angelino, Paolo; Riezman, Howard; Hatzimanikatis, Vassily

    2016-09-01

    We present a model-based method, designated Inverse Metabolic Control Analysis (IMCA), which can be used in conjunction with classical Metabolic Control Analysis for the analysis and design of cellular metabolism. We demonstrate the capabilities of the method by first developing a comprehensively curated kinetic model of sphingolipid biosynthesis in the yeast Saccharomyces cerevisiae. Next we apply IMCA using the model and integrating lipidomics data. The combinatorial complexity of the synthesis of sphingolipid molecules, along with the operational complexity of the participating enzymes of the pathway, presents an excellent case study for testing the capabilities of the IMCA. The exceptional agreement of the predictions of the method with genome-wide data highlights the importance and value of a comprehensive and consistent engineering approach for the development of such methods and models. Based on the analysis, we identified the class of enzymes regulating the distribution of sphingolipids among species and hydroxylation states, with the D-phospholipase SPO14 being one of the most prominent. The method and the applications presented here can be used for a broader, model-based inverse metabolic engineering approach.

  5. Preliminary study on mechanics-based rainfall kinetic energy

    Directory of Open Access Journals (Sweden)

    Yuan Jiuqin Ms.

    2014-09-01

    Full Text Available A raindrop impact power observation system was employed to observe the real-time raindrop impact power during a rainfall event and to analyze the corresponding rainfall characteristics. The experiments were conducted at different simulated rainfall intensities. As rainfall intensity increased, the observed impact power increased linearly indicating the power observation system would be satisfactory for characterizing rainfall erosivity. Momentum is the product of mass and velocity (Momentum=MV, which is related to the observed impact power value. Since there is no significant difference between momentum and impact power, observed impact power can represent momentum for different rainfall intensities. The relationship between momentum and the observed impact power provides a convenient way to calculate rainfall kinetic energy. The value of rainfall kinetic energy based on the observed impact power was higher than the classic rainfall kinetic energy. The rainfall impact power based kinetic energy and the classic rainfall kinetic energy showed linear correlation, which indicates that the raindrop impact power observation system can characterize rainfall kinetic energy. The article establishes a preliminary way to calculate rainfall kinetic energy by using the real-time observed momentum, providing a foundation for replacing the traditional methods for estimating kinetic energy of rainstorms.

  6. Enzyme inhibition by iminosugars

    DEFF Research Database (Denmark)

    López, Óscar; Qing, Feng-Ling; Pedersen, Christian Marcus

    2013-01-01

    Imino- and azasugar glycosidase inhibitors display pH dependant inhibition reflecting that both the inhibitor and the enzyme active site have groups that change protonation state with pH. With the enzyme having two acidic groups and the inhibitor one basic group, enzyme-inhibitor complexes...

  7. Effective use of enzyme microreactors : thermal, kinetic and ethical guidelines

    NARCIS (Netherlands)

    Swarts, J.W.

    2009-01-01

    Microreactor technology is reported to have many benefits over regular chemical methods. Due to the small dimensions over which temperature and concentration gradients can exist, mass and heat transfer can be very quick. This could minimize the time needed for heating and mixing, due to a reduction

  8. Kinetic studies of novel inhibitors of endomorphin degrading enzymes

    OpenAIRE

    Perlikowska, Renata; Fichna, Jakub; do-Rego, Jean Claude; Gach, Katarzyna; Janecka, Anna

    2011-01-01

    Endomorphins (EMs), two endogenous μ-opioid receptor selective ligands, are attractive lead compounds for opioid-based pain management studies. However, these peptides are quickly degraded by peptidases, in particular by dipeptidylpeptidase IV (DPP IV) and aminopeptidase M (APM). Targeting enzymatic degradation is one approach to prolong endomorphin activity. In this study we characterized the action of two new inhibitors of similar to endomorphins structure, Tyr-Pro-Ala-NH2 (EMDB-2) and Tyr-...

  9. Resonance Van Hove Singularities in Wave Kinetics

    CERN Document Server

    Shi, Yi-Kang

    2015-01-01

    Wave kinetic theory has been developed to describe the statistical dynamics of weakly nonlinear, dispersive waves. However, we show that systems which are generally dispersive can have resonant sets of wave modes with identical group velocities, leading to a local breakdown of dispersivity. This shows up as a geometric singularity of the resonant manifold and possibly as an infinite phase measure in the collision integral. Such singularities occur widely for classical wave systems, including acoustical waves, Rossby waves, helical waves in rotating fluids, light waves in nonlinear optics and also in quantum transport, e.g. kinetics of electron-hole excitations (matter waves) in graphene. These singularities are the exact analogue of the critical points found by Van Hove in 1953 for phonon dispersion relations in crystals. The importance of these singularities in wave kinetics depends on the dimension of phase space $D=(N-2)d$ ($d$ physical space dimension, $N$ the number of waves in resonance) and the degree ...

  10. Sulfide toxicity kinetics of a uasb reactor

    Directory of Open Access Journals (Sweden)

    D. R. Paula Jr.

    2009-12-01

    Full Text Available The effect of sulfide toxicity on kinetic parameters of anaerobic organic matter removal in a UASB (up-flow anaerobic sludge blanket reactor is presented. Two lab-scale UASB reactors (10.5 L were operated continuously during 12 months. The reactors were fed with synthetic wastes prepared daily using glucose, ammonium acetate, methanol and nutrient solution. One of the reactors also received increasing concentrations of sodium sulfide. For both reactors, the flow rate of 16 L.d-1 was held constant throughout the experiment, corresponding to a hydraulic retention time of 15.6 hours. The classic model for non-competitive sulfide inhibition was applied to the experimental data for determining the overall kinetic parameter of specific substrate utilization (q and the sulfide inhibition coefficient (Ki. The application of the kinetic parameters determined allows prediction of methanogenesis inhibition and thus the adoption of operating parameters to minimize sulfide toxicity in UASB reactors.

  11. Teaching Classical Mechanics using Smartphones

    CERN Document Server

    Chevrier, Joel; Ledenmat, Simon; Bsiesy, Ahmad

    2012-01-01

    Using a personal computer and a smartphone, iMecaProf is a software that provides a complete teaching environment for practicals associated to a Classical Mechanics course. iMecaProf proposes a visual, real time and interactive representation of data transmitted by a smartphone using the formalism of Classical Mechanics. Using smartphones is more than using a set of sensors. iMecaProf shows students that important concepts of physics they here learn, are necessary to control daily life smartphone operations. This is practical introduction to mechanical microsensors that are nowadays a key technology in advanced trajectory control. First version of iMecaProf can be freely downloaded. It will be tested this academic year in Universit\\'e Joseph Fourier (Grenoble, France)

  12. Classical Equations for Quantum Systems

    CERN Document Server

    Gell-Mann, Murray; Gell-Mann, Murray; Hartle, James B.

    1993-01-01

    The origin of the phenomenological deterministic laws that approximately govern the quasiclassical domain of familiar experience is considered in the context of the quantum mechanics of closed systems such as the universe as a whole. We investigate the requirements for coarse grainings to yield decoherent sets of histories that are quasiclassical, i.e. such that the individual histories obey, with high probability, effective classical equations of motion interrupted continually by small fluctuations and occasionally by large ones. We discuss these requirements generally but study them specifically for coarse grainings of the type that follows a distinguished subset of a complete set of variables while ignoring the rest. More coarse graining is needed to achieve decoherence than would be suggested by naive arguments based on the uncertainty principle. Even coarser graining is required in the distinguished variables for them to have the necessary inertia to approach classical predictability in the presence of t...

  13. Classical databases and knowledge organization

    DEFF Research Database (Denmark)

    Hjørland, Birger

    2015-01-01

    This paper considers classical bibliographic databases based on the Boolean retrieval model (such as MEDLINE and PsycInfo). This model is challenged by modern search engines and information retrieval (IR) researchers, who often consider Boolean retrieval a less efficient approach. The paper...... examines this claim and argues for the continued value of Boolean systems, which suggests two further considerations: (1) the important role of human expertise in searching (expert searchers and “information literate” users) and (2) the role of library and information science and knowledge organization (KO......) in the design and use of classical databases. An underlying issue is the kind of retrieval system for which one should aim. Warner’s (2010) differentiation between the computer science traditions and an older library-oriented tradition seems important; the former aim to transform queries automatically...

  14. From classical to quantum physics

    CERN Document Server

    Stehle, Philip

    2017-01-01

    Suitable for lay readers as well as students, this absorbing survey explores the twentieth-century transition from classical to quantum physics. Author Philip Stehle traces the shift in the scientific worldview from the work of Galileo, Newton, and Darwin to the modern-day achievements of Max Planck, Albert Einstein, Ernest Rutherford, Niels Bohr, and others of their generation. His insightful overview examines not only the history of quantum physics but also the ways that progress in the discipline changed our understanding of the physical world and forces of nature. This chronicle of the second revolution in the physical sciences conveys the excitement and suspense that new developments produced in the scientific community. The narrative ranges from the classical physics of the seventeenth-century to the emergence of quantum mechanics with the entrance of the electron, the rise of relativity theory, the development of atomic theory, and the recognition of wave-particle duality. Relevant mathematical details...

  15. A Companion to Classical Receptions

    Directory of Open Access Journals (Sweden)

    A. De Villiers

    2012-03-01

    Full Text Available This recent addition to the excellent Blackwell Companions series looks at the various forms of classical reception currently being researched as well as those deemed to have future importance. The diversity and volume of the themes and approaches contained in this book are truly impressive. As Hardwick and Stray state in their introduction, this collection “has been constructed on the basis that the activators of reception are many and varied and that we all gain from encountering examples from outside our own immediate areas of knowledge” (p. 4. Throughout the book they stay true to this motto and traditional approaches to classical reception are not given prominence over more recent (sometimes contentious approaches such as film studies, cultural politics and photography. The same goes for the various cultures involved and there is even a chapter on Greek drama in South Africa.

  16. Quantum Models of Classical World

    Directory of Open Access Journals (Sweden)

    Petr Hájíček

    2013-02-01

    Full Text Available This paper is a review of our recent work on three notorious problems of non-relativistic quantum mechanics: realist interpretation, quantum theory of classical properties, and the problem of quantum measurement. A considerable progress has been achieved, based on four distinct new ideas. First, objective properties are associated with states rather than with values of observables. Second, all classical properties are selected properties of certain high entropy quantum states of macroscopic systems. Third, registration of a quantum system is strongly disturbed by systems of the same type in the environment. Fourth, detectors must be distinguished from ancillas and the states of registered systems are partially dissipated and lost in the detectors. The paper has two aims: a clear explanation of all new results and a coherent and contradiction-free account of the whole quantum mechanics including all necessary changes of its current textbook version.

  17. Classical Corrections in String Cosmology

    CERN Document Server

    Brustein, Ram; Brustein, Ram; Madden, Richard

    1999-01-01

    An important element in a model of non-singular string cosmology is a phase in which classical corrections saturate the growth of curvature in a deSitter-like phase with a linearly growing dilaton (an `algebraic fixed point'). As the form of the classical corrections is not well known, here we look for evidence, based on a suggested symmetry of the action, scale factor duality and on conformal field theory considerations, that they can produce this saturation. It has previously been observed that imposing scale factor duality on the $O(\\alpha')$ corrections is not compatible with fixed point behavior. Here we present arguments that these problems persist to all orders in $\\alpha'$. We also present evidence for the form of a solution to the equations of motion using conformal perturbation theory, examine its implications for the form of the effective action and find novel fixed point structure.

  18. Potential Theory in Classical Electrodynamics

    CERN Document Server

    Engelhardt, Wolfgang

    2012-01-01

    In Maxwell's classical theory of electrodynamics the fields are frequently expressed by potentials in order to facilitate the solution of the first order system of equations. This method obscures, however, that there exists an inconsistency between Faraday's law of induction and Maxwell's flux law. As a consequence of this internal contradiction there is neither gauge invariance, nor exist unique solutions in general. It is also demonstrated that inhomogeneous wave equations cannot be solved by retarded integrals.

  19. Semi-classical signal analysis

    CERN Document Server

    Laleg-Kirati, Taous-Meriem; Sorine, Michel

    2010-01-01

    This study introduces a new signal analysis method called SCSA, based on a semi-classical approach. The main idea in the SCSA is to interpret a pulse-shaped signal as a potential of a Schr\\"odinger operator and then to use the discrete spectrum of this operator for the analysis of the signal. We present some numerical examples and the first results obtained with this method on the analysis of arterial blood pressure waveforms.

  20. Quantum manifolds with classical limit

    CERN Document Server

    Hohmann, Manuel; Wohlfarth, Mattias N R

    2008-01-01

    We propose a mathematical model of quantum spacetime as an infinite-dimensional manifold locally homeomorphic to an appropriate Schwartz space. This extends and unifies both the standard function space construction of quantum mechanics and the manifold structure of spacetime. In this picture we demonstrate that classical spacetime emerges as a finite-dimensional manifold through the topological identification of all quantum points with identical position expectation value. We speculate on the possible relevance of this geometry to quantum field theory and gravity.

  1. Semi-classical signal analysis

    KAUST Repository

    Laleg-Kirati, Taous-Meriem

    2012-09-30

    This study introduces a new signal analysis method, based on a semi-classical approach. The main idea in this method is to interpret a pulse-shaped signal as a potential of a Schrödinger operator and then to use the discrete spectrum of this operator for the analysis of the signal. We present some numerical examples and the first results obtained with this method on the analysis of arterial blood pressure waveforms. © 2012 Springer-Verlag London Limited.

  2. Nanomaterials with enzyme-like characteristics (nanozymes): next-generation artificial enzymes.

    Science.gov (United States)

    Wei, Hui; Wang, Erkang

    2013-07-21

    Over the past few decades, researchers have established artificial enzymes as highly stable and low-cost alternatives to natural enzymes in a wide range of applications. A variety of materials including cyclodextrins, metal complexes, porphyrins, polymers, dendrimers and biomolecules have been extensively explored to mimic the structures and functions of naturally occurring enzymes. Recently, some nanomaterials have been found to exhibit unexpected enzyme-like activities, and great advances have been made in this area due to the tremendous progress in nano-research and the unique characteristics of nanomaterials. To highlight the progress in the field of nanomaterial-based artificial enzymes (nanozymes), this review discusses various nanomaterials that have been explored to mimic different kinds of enzymes. We cover their kinetics, mechanisms and applications in numerous fields, from biosensing and immunoassays, to stem cell growth and pollutant removal. We also summarize several approaches to tune the activities of nanozymes. Finally, we make comparisons between nanozymes and other catalytic materials (other artificial enzymes, natural enzymes, organic catalysts and nanomaterial-based catalysts) and address the current challenges and future directions (302 references).

  3. A Simple Classroom Teaching Technique to Help Students Understand Michaelis-Menten Kinetics

    Science.gov (United States)

    Runge, Steven W.; Hill, Brent J. F.; Moran, William M.

    2006-01-01

    A new, simple classroom technique helps cell biology students understand principles of Michaelis-Menten enzyme kinetics. A student mimics the enzyme and the student's hand represents the enzyme's active site. The catalytic event is the transfer of marbles (substrate molecules) by hand from one plastic container to another. As predicted, increases…

  4. Application of artificial neural networks and DFT-based parameters for prediction of reaction kinetics of ethylbenzene dehydrogenase

    Science.gov (United States)

    Szaleniec, Maciej; Witko, Małgorzata; Tadeusiewicz, Ryszard; Goclon, Jakub

    2006-03-01

    Artificial neural networks (ANNs) are used for classification and prediction of enzymatic activity of ethylbenzene dehydrogenase from EbN1 Azoarcus sp. bacterium. Ethylbenzene dehydrogenase (EBDH) catalyzes stereo-specific oxidation of ethylbenzene and its derivates to alcohols, which find its application as building blocks in pharmaceutical industry. ANN systems are trained based on theoretical variables derived from Density Functional Theory (DFT) modeling, topological descriptors, and kinetic parameters measured with developed spectrophotometric assay. Obtained models exhibit high degree of accuracy (100% of correct classifications, correlation between predicted and experimental values of reaction rates on the 0.97 level). The applicability of ANNs is demonstrated as useful tool for the prediction of biochemical enzyme activity of new substrates basing only on quantum chemical calculations and simple structural characteristics. Multi Linear Regression and Molecular Field Analysis (MFA) are used in order to compare robustness of ANN and both classical and 3D-quantitative structure-activity relationship (QSAR) approaches.

  5. Non-Isothermal Kinetics.

    Science.gov (United States)

    Brown, M. E.; Phillpotts, C. A. R.

    1978-01-01

    Discusses the principle of nonisothermal kinetics and some of the factors involved in such reactions, especially when considering the reliability of the kinetic parameters, compared to those of isothermal conditions. (GA)

  6. Quantum-Classical Correspondence: Dynamical Quantization and the Classical Limit

    Energy Technology Data Exchange (ETDEWEB)

    Turner, L [Los Alamos National Laboratory, Los Alamos, NM 87545 (United States)

    2004-11-12

    In only 150 pages, not counting appendices, references, or the index, this book is one author's perspective of the massive theoretical and philosophical hurdles in the no-man's-land separating the classical and quantum domains of physics. It ends with him emphasizing his own theoretical contribution to this area. In his own words, he has attempted to answer: 1. How can we obtain the quantum dynamics of open systems initially described by the equations of motion of classical physics (quantization process) 2. How can we retrieve classical dynamics from the quantum mechanical equations of motion by means of a classical limiting process (dequantization process). However, this monograph seems overly ambitious. Although the publisher's description refers to this book as an accessible entre, we find that this author scrambles too hastily over the peaks of information that are contained in his large collection of 272 references. Introductory motivating discussions are lacking. Profound ideas are glossed over superficially and shoddily. Equations morph. But no new convincing understanding of the physical world results. The author takes the viewpoint that physical systems are always in interaction with their environment and are thus not isolated and, therefore, not Hamiltonian. This impels him to produce a method of quantization of these stochastic systems without the need of a Hamiltonian. He also has interest in obtaining the classical limit of the quantized results. However, this reviewer does not understand why one needs to consider open systems to understand quantum-classical correspondence. The author demonstrates his method using various examples of the Smoluchowski form of the Fokker--Planck equation. He then renders these equations in a Wigner representation, uses what he terms an infinitesimality condition, and associates with a constant having the dimensions of an action. He thereby claims to develop master equations, such as the Caldeira

  7. Quantum Transitions Between Classical Histories: Bouncing Cosmologies

    CERN Document Server

    Hartle, James

    2015-01-01

    In a quantum theory of gravity spacetime behaves classically when quantum probabilities are high for histories of geometry and field that are correlated in time by the Einstein equation. Probabilities follow from the quantum state. This quantum perspective on classicality has important implications: (a) Classical histories are generally available only in limited patches of the configuration space on which the state lives. (b) In a given patch states generally predict relative probabilities for an ensemble of possible classical histories. (c) In between patches classical predictability breaks down and is replaced by quantum evolution connecting classical histories in different patches. (d) Classical predictability can break down on scales well below the Planck scale, and with no breakdown in the classical equations of motion. We support and illustrate (a)-(d) by calculating the quantum transition across the de Sitter like throat connecting asymptotically classical, inflating histories in the no-boundary quantu...

  8. Classical and quantum dynamics from classical paths to path integrals

    CERN Document Server

    Dittrich, Walter

    2016-01-01

    Graduate students who want to become familiar with advanced computational strategies in classical and quantum dynamics will find here both the fundamentals of a standard course and a detailed treatment of the time-dependent oscillator, Chern-Simons mechanics, the Maslov anomaly and the Berry phase, to name a few. Well-chosen and detailed examples illustrate the perturbation theory, canonical transformations, the action principle and demonstrate the usage of path integrals. This new edition has been revised and enlarged with chapters on quantum electrodynamics, high energy physics, Green’s functions and strong interaction.

  9. Intuitionism vs. classicism a mathematical attack on classical logic

    CERN Document Server

    Haverkamp, Nick

    2015-01-01

    In the early twentieth century, the Dutch mathematician L.E.J. Brouwer launched a powerful attack on the prevailing mathematical methods and theories. He developed a new kind of constructive mathematics, called intuitionism, which seems to allow for a rigorous refutation of widely accepted mathematical assumptions including fundamental principles of classical logic. Following an intense mathematical debate esp. in the 1920s, Brouwer's revolutionary criticism became a central philosophical concern in the 1970s, when Michael Dummett tried to substantiate it with meaning-theoretic considerations.

  10. Activity assessment of microbial fibrinolytic enzymes.

    Science.gov (United States)

    Kotb, Essam

    2013-08-01

    Conversion of fibrinogen to fibrin inside blood vessels results in thrombosis, leading to myocardial infarction and other cardiovascular diseases. In general, there are four therapy options: surgical operation, intake of antiplatelets, anticoagulants, or fibrinolytic enzymes. Microbial fibrinolytic enzymes have attracted much more attention than typical thrombolytic agents because of the expensive prices and the side effects of the latter. The fibrinolytic enzymes were successively discovered from different microorganisms, the most important among which is the genus Bacillus. Microbial fibrinolytic enzymes, especially those from food-grade microorganisms, have the potential to be developed as functional food additives and drugs to prevent or cure thrombosis and other related diseases. There are several assay methods for these enzymes; this may due to the insolubility of substrate, fibrin. Existing assay methods can be divided into three major groups. The first group consists of assay of fibrinolytic activity with natural proteins as substrates, e.g., fibrin plate methods. The second and third groups of assays are suitable for kinetic studies and are based on the determination of hydrolysis of synthetic peptide esters. This review will deal primarily with the microorganisms that have been reported in literature to produce fibrinolytic enzymes and the first review discussing the methods used to assay the fibrinolytic activity.

  11. Cloning and functional analysis of a high DP fructan:fructan 1-fructosyl transferase from Echinops ritro (Asteraceae): comparison of the native and recombinant enzymes.

    Science.gov (United States)

    Van den Ende, Wim; Clerens, Stefan; Vergauwen, Rudy; Boogaerts, David; Le Roy, Katrien; Arckens, Lutgarde; Van Laere, André

    2006-01-01

    Inulin-type fructans are the simplest and most studied fructans and have become increasingly popular as prebiotic health-improving compounds. A natural variation in the degree of polymerization (DP) of inulins is observed within the family of the Asteraceae. Globe thistle (Echinops ritro), artichoke (Cynara scolymus), and Viguiera discolor biosynthesize fructans with a considerably higher DP than Cichorium intybus (chicory), Helianthus tuberosus (Jerusalem artichoke), and Dahlia variabilis. The higher DP in some species can be explained by the presence of special fructan:fructan 1-fructosyl transferases (high DP 1-FFTs), different from the classical low DP 1-FFTs. Here, the RT-PCR-based cloning of a high DP 1-FFT cDNA from Echinops ritro is described, starting from peptide sequence information derived from the purified native high DP 1-FFT enzyme. The cDNA was successfully expressed in Pichia pastoris. A comparison is made between the mass fingerprints of the native, heterodimeric enzyme and its recombinant, monomeric counterpart (mass fingerprints and kinetical analysis) showing that they have very similar properties. The recombinant enzyme is a functional 1-FFT lacking invertase and 1-SST activities, but shows a small intrinsic 1-FEH activity. The enzyme is capable of producing a high DP inulin pattern in vitro, similar to the one observed in vivo. Depending on conditions, the enzyme is able to produce fructo-oligosaccharides (FOS) as well. Therefore, the enzyme might be suitable for both FOS and high DP inulin production in bioreactors. Alternatively, introduction of the high DP 1-FFT gene in chicory, a crop widely used for inulin extraction, could lead to an increase in DP which is useful for a number of specific industrial applications. 1-FFT expression analysis correlates well with high DP fructan accumulation in vivo, suggesting that the enzyme is responsible for high DP fructan formation in planta.

  12. Methods of nonlinear kinetics

    OpenAIRE

    Gorban, A. N.; Karlin, I.V.

    2003-01-01

    Nonlinear kinetic equations are reviewed for a wide audience of specialists and postgraduate students in physics, mathematical physics, material science, chemical engineering and interdisciplinary research. Contents: The Boltzmann equation, Phenomenology and Quasi-chemical representation of the Boltzmann equation, Kinetic models, Discrete velocity models, Direct simulation, Lattice Gas and Lattice Boltzmann models, Minimal Boltzmann models for flows at low Knudsen number, Other kinetic equati...

  13. The Relation between Classical and Quantum Electrodynamics

    Directory of Open Access Journals (Sweden)

    Mario Bacelar Valente

    2011-01-01

    Full Text Available Quantum electrodynamics presents intrinsic limitations in the description of physical processes that make it impossible to recover from it the type of description we have in classical electrodynamics. Hence one cannot consider classical electrodynamics as reducing to quantum electrodynamics and being recovered from it by some sort of limiting procedure. Quantum electrodynamics has to be seen not as an more fundamental theory, but as an upgrade of classical electrodynamics, which permits an extension of classical theory to the description of phenomena that, while being related to the conceptual framework of the classical theory, cannot be addressed from the classical theory.

  14. Marine Microbiological Enzymes: Studies with Multiple Strategies and Prospects.

    Science.gov (United States)

    Wang, Yan; Song, Qinghao; Zhang, Xiao-Hua

    2016-09-22

    Marine microorganisms produce a series of promising enzymes that have been widely used or are potentially valuable for our daily life. Both classic and newly developed biochemistry technologies have been broadly used to study marine and terrestrial microbiological enzymes. In this brief review, we provide a research update and prospects regarding regulatory mechanisms and related strategies of acyl-homoserine lactones (AHL) lactonase, which is an important but largely unexplored enzyme. We also detail the status and catalytic mechanism of the main types of polysaccharide-degrading enzymes that broadly exist among marine microorganisms but have been poorly explored. In order to facilitate understanding, the regulatory and synthetic biology strategies of terrestrial microorganisms are also mentioned in comparison. We anticipate that this review will provide an outline of multiple strategies for promising marine microbial enzymes and open new avenues for the exploration, engineering and application of various enzymes.

  15. Marine Microbiological Enzymes: Studies with Multiple Strategies and Prospects

    Directory of Open Access Journals (Sweden)

    Yan Wang

    2016-09-01

    Full Text Available Marine microorganisms produce a series of promising enzymes that have been widely used or are potentially valuable for our daily life. Both classic and newly developed biochemistry technologies have been broadly used to study marine and terrestrial microbiological enzymes. In this brief review, we provide a research update and prospects regarding regulatory mechanisms and related strategies of acyl-homoserine lactones (AHL lactonase, which is an important but largely unexplored enzyme. We also detail the status and catalytic mechanism of the main types of polysaccharide-degrading enzymes that broadly exist among marine microorganisms but have been poorly explored. In order to facilitate understanding, the regulatory and synthetic biology strategies of terrestrial microorganisms are also mentioned in comparison. We anticipate that this review will provide an outline of multiple strategies for promising marine microbial enzymes and open new avenues for the exploration, engineering and application of various enzymes.

  16. Kinetic resolution and chemoenzymatic dynamic kinetic resolution of functionalized gamma-hydroxy amides.

    Science.gov (United States)

    Fransson, Ann-Britt L; Borén, Linnéa; Pàmies, Oscar; Bäckvall, Jan-E

    2005-04-01

    [reaction: see text] An efficient kinetic resolution of racemic gamma-hydroxy amides 1 was performed via Pseudomas cepacia lipase (PS-C)-catalyzed transesterification. The enzyme PS-C tolerates both variation in the chain length and different functionalities giving good to high enantioselectivity (E values of up to >250). The combination of enzymatic kinetic resolution with a ruthenium-catalyzed racemization led to a dynamic kinetic resolution. The use of 2,4-dimethyl-3-pentanol as a hydrogen source to suppress ketone formation in the dynamic kinetic resolution yields the corresponding acetates in good yield and good to high enantioselectivity (ee's up to 98%). The synthetic utility of this procedure was illustrated by the practical synthesis of the versatile intermediate gamma-lactone (R)-5-methyltetrahydrofuran-2-one.

  17. Microtubule dynamics. II. Kinetics of self-assembly

    DEFF Research Database (Denmark)

    Flyvbjerg, H.; Jobs, E.

    1997-01-01

    Inverse scattering theory describes the conditions necessary and sufficient to determine an unknown potential from known scattering data. No similar theory exists for when and how one may deduce the kinetics of an unknown chemical reaction from quantitative information about its final state and its...... various initial concentrations of monomers. Distinct mathematical properties of the kinetics of the unknown reaction pathway are found. These properties are shown to restrict the kinetics to a single model that generalizes Oosawa's classical nucleation-polymerization model. The methods used here...

  18. Classical Optics and its Applications

    Science.gov (United States)

    Mansuripur, Masud

    2009-02-01

    Preface; Introduction; 1. Abbe's sine condition; 2. Fourier optics; 3. Effect of polarization on diffraction in systems of high numerical aperture; 4. Gaussian beam optics; 5. Coherent and incoherent imaging; 6. First-order temporal coherence in classical optics; 7. The Van Cittert-Zernike theorem; 8. Partial polarization, Stokes parameters, and the Poincarè Sphere; 9. Second-order coherence and the Hanbury Brown - Twiss experiment; 10. What in the world are surface plasmons?; 11. Surface plasmon polaritons on metallic surfaces; 12. The Faraday effecy; 13. The magneto-optical Kerr effect; 14. The Sagnac interferometer; 15. Fabry-Perot etalons in polarized light; 16. The Ewald-Oseen extinction theorem; 17. Reciprocity in classical Linear optics; 18. Optical pulse compression; 19. The uncertainty principle in classical optics; 20. Omni-directional dielectric mirrors; 21. Optical vortices; 22. Geometric-optical rays, Poynting's vector, and field momenta; 23. Doppler shift, stellar aberration, and convection of light by moving Media; 24. Diffraction gratings; 25. Diffractive optical elements; 26. The talbot effect; 27. Some quirks of total internal reflection; 28. Evanescent coupling; 29. Internal and external conical refraction; 30. Transmission of light through small elliptical apertures; 31. The method of Fox and Li; 32. The beam propagation method; 33. Launching light into a Fiber; 34. The optics of demiconductor fiode Laser; 35. Michelson's dtellar interferometer; 36. Bracewell's interferometric telescope; 37. Scanning optical microscopy; 38. Zernike's method of phase contrast; 39. Polarization microscopy; 40. Nomarski's differential interference contrast microscope; 41. The Van Leeuwenhoek microscope; 42. Projection photolithography; 43. Interaction of light with subwavelength structures; 44 The Ronchi test; 45. The Shack-Hartmann Wavefront sensor; 46. Ellipsometry; 47. Holography and holographic interferometry; 48. Self-focusing in non-linear optical media; 49

  19. Classical trajectories and quantum tunneling

    CERN Document Server

    Ivlev, B I

    2003-01-01

    The problem of inter-band tunneling in a semiconductor (Zener breakdown) in a nonstationary and homogeneous electric field is solved exactly. Using the exact analytical solution, the approximation based on classical trajectories is studied. A new mechanism of enhanced tunneling through static non-one-dimensional barriers is proposed in addition to well known normal tunneling solely described by a trajectory in imaginary time. Under certain conditions on the barrier shape and the particle energy, the probability of enhanced tunneling is not exponentially small even for non-transparent barriers, in contrast to the case of normal tunneling.

  20. Motions of Classical Charged Tachyons

    CERN Document Server

    Davidson, M P

    2001-01-01

    It is shown by numerical simulation that classical charged tachyons have self-orbiting helical solutions in a narrow neighborhood of certain discrete values for the velocity when the electromagnetic interaction is described by Feynman-Wheeler electrodynamics. The force rapidly oscillates between attractive and repulsive as a function of velocity in this neighborhood. Causal electrodynamics is also considered, and in this case it is found that when the force is attractive the tachyon loses energy to radiation. Only certain narrow ranges of velocity give attractive forces, and a geometrical derivation of these special velocities is given. Possible implications of these results for hidden variable theories of quantum mechanics are conjectured.

  1. Solar Activity and Classical Physics

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This review of solar physics emphasizes several of the more conspicuous scientific puzzles posed by contemporary observational knowledge of the magnetic activity of the Sun. The puzzles emphasize how much classical physics we have yet to learn from the Sun. The physics of solar activity is based on the principles of Newton, Maxwell, Lorentz, Boltzmann, et. al., along with the principles of radiative transfer. In the large, these principles are expressed by magnetohydrodynamics. A brief derivation of the magnetohydrodynamic induction and momentum equations is provided, with a discussion of popular misconceptions.

  2. Classical databases and knowledge organization

    DEFF Research Database (Denmark)

    Hjørland, Birger

    2015-01-01

    examines this claim and argues for the continued value of Boolean systems, which suggests two further considerations: (1) the important role of human expertise in searching (expert searchers and “information literate” users) and (2) the role of library and information science and knowledge organization (KO......) in the design and use of classical databases. An underlying issue is the kind of retrieval system for which one should aim. Warner’s (2010) differentiation between the computer science traditions and an older library-oriented tradition seems important; the former aim to transform queries automatically...

  3. Lectures on classical differential geometry

    CERN Document Server

    Struik, Dirk J

    1988-01-01

    Elementary, yet authoritative and scholarly, this book offers an excellent brief introduction to the classical theory of differential geometry. It is aimed at advanced undergraduate and graduate students who will find it not only highly readable but replete with illustrations carefully selected to help stimulate the student's visual understanding of geometry. The text features an abundance of problems, most of which are simple enough for class use, and often convey an interesting geometrical fact. A selection of more difficult problems has been included to challenge the ambitious student.Writ

  4. Manipulating single enzymes by an external harmonic force

    DEFF Research Database (Denmark)

    Lomholt, Michael A; Urbakh, Michael; Metzler, Ralf;

    2007-01-01

    We study a Michaelis-Menten reaction for a single two-state enzyme molecule, whose transition rates between the two conformations are modulated by an harmonically oscillating external force. In particular, we obtain a range of optimal driving frequencies for changing the conformation of the enzyme......, thereby controlling the enzymatic activity (i.e., product formation). This analysis demonstrates that it is, in principle, possible to obtain information about particular rates within the kinetic scheme....

  5. Supramolecular Tectonics for Enzyme-like Reagents Ⅱ Mimic and Assay of the Enzyme Containing Fe(Ⅱ, Ⅲ)

    Institute of Scientific and Technical Information of China (English)

    MAO LuYuan; CAI RuXiu; HUANG Bo; ZOU GuoLin; ZHOU Hui; GAO ChengZhuo

    2001-01-01

    @@ The enzyme-likes and bioactive species were closely related with the life phenomena and served as the reagent of bioassy1,2. In present works, the flow cytometry (FCM) and rapid-scanning stopped-flow (RSSF) spectroscopy combine with the stopped-flow difference UV/Vis spectra, FT-IR and other methods of assay, being used to study the biomimetic reaction and enzyme mimic. Based on catalytic kinetics of enzyme reaction3,4, the reaction mechanisms of the enzyme-likes had been studied and some new methods of kinetic determination were proposed. The study and methods not only provided the basic theoretical models for the life science, but also widened the application fields of biomimetic and analytical chemistry. The main contents of our works and the supramolecular models can be described as follows:

  6. Classical and Quantum Thermal Physics

    Science.gov (United States)

    Prasad, R.

    2016-11-01

    List of figures; List of tables; Preface; Acknowledgement; Dedication; 1. The kinetic theory of gases; 2. Ideal to real gas, viscosity, conductivity and diffusion; 3. Thermodynamics: definitions and Zeroth law; 4. First Law of Thermodynamics and some of its applications; 5. Second Law of Thermodynamics and some of its applications; 6. TdS equations and their applications; 7. Thermodynamic functions, potentials, Maxwell equations, the Third Law and equilibrium; 8. Some applications of thermodynamics to problems of physics and engineering; 9. Application of thermodynamics to chemical reactions; 10. Quantum thermodynamics; 11. Some applications of quantum thermodynamics; 12. Introduction to the thermodynamics of irreversible processes; Index.

  7. The revision of classical stock model

    Institute of Scientific and Technical Information of China (English)

    YE Bai-qing; WANG Hong-li

    2001-01-01

    On the basis of the analysis of classical stock model, according to th e limitation of the model, the article puts forward the revision of classical mo del and enforces the applicability of the stock model.

  8. The revision of classical stock model

    Institute of Scientific and Technical Information of China (English)

    叶柏青; 王洪利

    2001-01-01

    On the basis of the analysis of classical stock model, according to the limitation of the model, the article puts forward the revision of classical model and enforces the applicability of the stock model.

  9. A model study of sequential enzyme reactions and electrostatic channeling.

    Science.gov (United States)

    Eun, Changsun; Kekenes-Huskey, Peter M; Metzger, Vincent T; McCammon, J Andrew

    2014-03-14

    We study models of two sequential enzyme-catalyzed reactions as a basic functional building block for coupled biochemical networks. We investigate the influence of enzyme distributions and long-range molecular interactions on reaction kinetics, which have been exploited in biological systems to maximize metabolic efficiency and signaling effects. Specifically, we examine how the maximal rate of product generation in a series of sequential reactions is dependent on the enzyme distribution and the electrostatic composition of its participant enzymes and substrates. We find that close proximity between enzymes does not guarantee optimal reaction rates, as the benefit of decreasing enzyme separation is countered by the volume excluded by adjacent enzymes. We further quantify the extent to which the electrostatic potential increases the efficiency of transferring substrate between enzymes, which supports the existence of electrostatic channeling in nature. Here, a major finding is that the role of attractive electrostatic interactions in confining intermediate substrates in the vicinity of the enzymes can contribute more to net reactive throughput than the directional properties of the electrostatic fields. These findings shed light on the interplay of long-range interactions and enzyme distributions in coupled enzyme-catalyzed reactions, and their influence on signaling in biological systems.

  10. Enzymes for improved biomass conversion

    Science.gov (United States)

    Brunecky, Roman; Himmel, Michael E.

    2016-02-02

    Disclosed herein are enzymes and combinations of the enzymes useful for the hydrolysis of cellulose and the conversion of biomass. Methods of degrading cellulose and biomass using enzymes and cocktails of enzymes are also disclosed.

  11. Handling and Sensing of Single Enzyme Molecules: From Fluorescence Detection towards Nanoscale Electrical Measurements

    NARCIS (Netherlands)

    Mathwig, Klaus; Chi, Qijin; Lemay, Serge G.; Rassaei, Liza

    2016-01-01

    Classical methods to study single enzyme molecules have pro- vided valuable information about the distribution of conforma- tional heterogeneities, reaction mechanisms, and transients in enzymatic reactions when individual molecules instead of an averaging ensemble are studied. Here, we highlight ma

  12. Seven Steps Towards the Classical World

    OpenAIRE

    Allori, Valia; Dürr, Detlef; Goldstein, Shelly; Zanghí, Nino

    2001-01-01

    Classical physics is about real objects, like apples falling from trees, whose motion is governed by Newtonian laws. In standard Quantum Mechanics only the wave function or the results of measurements exist, and to answer the question of how the classical world can be part of the quantum world is a rather formidable task. However, this is not the case for Bohmian mechanics, which, like classical mechanics, is a theory about real objects. In Bohmian terms, the problem of the classical limit be...

  13. About the modern house - and the classical

    DEFF Research Database (Denmark)

    Hauberg, Jørgen

    2010-01-01

    In text and illustrations describes the classical house and the classical city, represented by Andrea Palladio (1508-80), and the modern house, the modern city and building task, represented by Le Corbusier (1857-1965).......In text and illustrations describes the classical house and the classical city, represented by Andrea Palladio (1508-80), and the modern house, the modern city and building task, represented by Le Corbusier (1857-1965)....

  14. Unhairing with enzymes

    OpenAIRE

    Crispim, A.; Mota, M.

    2003-01-01

    The use of enzymes in the leather industry is increasing and their application is being widened to include operations such as de-greasing, unhairing and other wet-end operations. Enzymes can also be used to assist with recycling leather wastes as well as to avoid pollution. The present work is devoted to illustrate the potential application of enzymes in unhairing without hair destruction. Enzymatic unhairing is based upon the weakening of the epidermis basal layer to which the hair is at...

  15. Overuse injuries in classical ballet.

    Science.gov (United States)

    Khan, K; Brown, J; Way, S; Vass, N; Crichton, K; Alexander, R; Baxter, A; Butler, M; Wark, J

    1995-05-01

    Successful management of classical ballet dancers with overuse injuries requires an understanding of the art form, precise knowledge of anatomy and awareness of certain conditions. Turnout is the single most fundamental physical attribute in classical ballet and 'forcing turnout' frequently contributes to overuse injuries. Common presenting conditions arising from the foot and ankle include problems at the first metatarsophalangeal joint, second metatarsal stress fractures, flexor hallucis longus tendinitis and anterior and posterior ankle impingement syndromes. Persistent shin pain in dancers is often due to chronic compartment syndrome, stress fracture of the posteromedial or anterior tibia. Knee pain can arise from patellofemoral syndrome, patellar tendon insertional pathologies, or a combination of both. Hip and back problems are also prevalent in dancers. To speed injury recovery of dancers, it is important for the sports medicine team to cooperate fully. This permits the dancer to benefit from accurate diagnosis, technique correction where necessary, the full range of manual therapies to joint and soft tissue, appropriate strengthening programmes and maintenance of dance fitness during any time out of class with Pilates-based exercises and nutrition advice. Most overuse ballet conditions respond well to a combination of conservative therapies. Those dancers that do require surgical management still depend heavily on ballet-specific rehabilitation for a complete recovery.

  16. Classical equations for quantum systems

    Energy Technology Data Exchange (ETDEWEB)

    Gell-Mann, M. (Theoretical Astrophysics Group (T-6), Los Alamos National Laboratory, Los Alamos, New Mexico 87545) (United States) (Santa Fe Institute, 1660 Old Pecos Trail, Santa Fe, New Mexico 87501); Hartle, J.B. (Department of Physics, University of California enSanta Barbara, Santa Barbara, (California) 93106)

    1993-04-15

    The origin of the phenomenological deterministic laws that approximately govern the quasiclassical domain of familiar experience is considered in the context of the quantum mechanics of closed systems such as the universe as a whole. A formulation of quantum mechanics is used that predicts probabilities for the individual members of a set of alternative coarse-grained histories that [ital decohere], which means that there is negligible quantum interference between the individual histories in the set. We investigate the requirements for coarse grainings to yield decoherent sets of histories that are quasiclassical, i.e., such that the individual histories obey, with high probability, effective classical equations of motion interrupted continually by small fluctuations and occasionally by large ones. We discuss these requirements generally but study them specifically for coarse grainings of the type that follows a distinguished subset of a complete set of variables while ignoring the rest. More coarse graining is needed to achieve decoherence than would be suggested by naive arguments based on the uncertainty principle. Even coarser graining is required in the distinguished variables for them to have the necessary inertia to approach classical predictability in the presence of the noise consisting of the fluctuations that typical mechanisms of decoherence produce. We describe the derivation of phenomenological equations of motion explicitly for a particular class of models.

  17. Physiological characteristics of classical ballet.

    Science.gov (United States)

    Schantz, P G; Astrand, P O

    1984-10-01

    The aerobic and anaerobic energy yield during professional training sessions ("classes") of classical ballet as well as during rehearsed and performed ballets has been studied by means of oxygen uptake, heart rate, and blood lactate concentration determinations on professional ballet dancers from the Royal Swedish Ballet in Stockholm. The measured oxygen uptake during six different normal classes at the theatre averaged about 35-45% of the maximal oxygen uptake, and the blood lactate concentration averaged 3 mM (N = 6). During 10 different solo parts of choreographed dance (median length = 1.8 min) representative for moderately to very strenuous dance, an average oxygen uptake (measured during the last minute) of 80% of maximum and blood lactate concentration of 10 mM was measured (N = 10). In addition, heart rate registrations from soloists in different ballets during performance and final rehearsals frequently indicated a high oxygen uptake relative to maximum and an average blood lactate concentration of 11 mM (N = 5). Maximal oxygen uptake, determined in 1971 (N = 11) and 1983 (N = 13) in two different groups of dancers, amounted to on the average 51 and 56 ml X min-1 X kg-1 for the females and males, respectively. In conclusion, classical ballet is a predominantly intermittent type of exercise. In choreographed dance each exercise period usually lasts only a few minutes, but can be very demanding energetically, while during the dancers' basic training sessions, the energy yield is low.

  18. Classical Black Holes Are Hot

    CERN Document Server

    Curiel, Erik

    2014-01-01

    In the early 1970s it is was realized that there is a striking formal analogy between the Laws of black-hole mechanics and the Laws of classical thermodynamics. Before the discovery of Hawking radiation, however, it was generally thought that the analogy was only formal, and did not reflect a deep connection between gravitational and thermodynamical phenomena. It is still commonly held that the surface gravity of a stationary black hole can be construed as a true physical temperature and its area as a true entropy only when quantum effects are taken into account; in the context of classical general relativity alone, one cannot cogently construe them so. Does the use of quantum field theory in curved spacetime offer the only hope for taking the analogy seriously? I think the answer is `no'. To attempt to justify that answer, I shall begin by arguing that the standard argument to the contrary is not physically well founded, and in any event begs the question. Looking at the various ways that the ideas of "tempe...

  19. Fluctuations in classical sum rules.

    Science.gov (United States)

    Elton, John R; Lakshminarayan, Arul; Tomsovic, Steven

    2010-10-01

    Classical sum rules arise in a wide variety of physical contexts. Asymptotic expressions have been derived for many of these sum rules in the limit of long orbital period (or large action). Although sum-rule convergence may well be exponentially rapid for chaotic systems in a global phase-space sense with time, individual contributions to the sums may fluctuate with a width which diverges in time. Our interest is in the global convergence of sum rules as well as their local fluctuations. It turns out that a simple version of a lazy baker map gives an ideal system in which classical sum rules, their corrections, and their fluctuations can be worked out analytically. This is worked out in detail for the Hannay-Ozorio sum rule. In this particular case the rate of convergence of the sum rule is found to be governed by the Pollicott-Ruelle resonances, and both local and global boundaries for which the sum rule may converge are given. In addition, the width of the fluctuations is considered and worked out analytically, and it is shown to have an interesting dependence on the location of the region over which the sum rule is applied. It is also found that as the region of application is decreased in size the fluctuations grow. This suggests a way of controlling the length scale of the fluctuations by considering a time dependent phase-space volume, which for the lazy baker map decreases exponentially rapidly with time.

  20. Simplification of the unified gas kinetic scheme

    Science.gov (United States)

    Chen, Songze; Guo, Zhaoli; Xu, Kun

    2016-08-01

    The unified gas kinetic scheme (UGKS) is an asymptotic preserving (AP) scheme for kinetic equations. It is superior for transition flow simulation and has been validated in the past years. However, compared to the well-known discrete ordinate method (DOM), which is a classical numerical method solving the kinetic equations, the UGKS needs more computational resources. In this study, we propose a simplification of the unified gas kinetic scheme. It allows almost identical numerical cost as the DOM, but predicts numerical results as accurate as the UGKS. In the simplified scheme, the numerical flux for the velocity distribution function and the numerical flux for the macroscopic conservative quantities are evaluated separately. The equilibrium part of the UGKS flux is calculated by analytical solution instead of the numerical quadrature in velocity space. The simplification is equivalent to a flux hybridization of the gas kinetic scheme for the Navier-Stokes (NS) equations and the conventional discrete ordinate method. Several simplification strategies are tested, through which we can identify the key ingredient of the Navier-Stokes asymptotic preserving property. Numerical tests show that, as long as the collision effect is built into the macroscopic numerical flux, the numerical scheme is Navier-Stokes asymptotic preserving, regardless the accuracy of the microscopic numerical flux for the velocity distribution function.