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Sample records for affine toda model

  1. ODE/IM correspondence and modified affine Toda field equations

    NASA Astrophysics Data System (ADS)

    Ito, Katsushi; Locke, Christopher

    2014-08-01

    We study the two-dimensional affine Toda field equations for affine Lie algebra gˆ modified by a conformal transformation and the associated linear equations. In the conformal limit, the associated linear problem reduces to a (pseudo-)differential equation. For classical affine Lie algebra gˆ, we obtain a (pseudo-)differential equation corresponding to the Bethe equations for the Langlands dual of the Lie algebra g, which were found by Dorey et al. in study of the ODE/IM correspondence.

  2. ODE/IM correspondence and Bethe ansatz for affine Toda field equations

    NASA Astrophysics Data System (ADS)

    Ito, Katsushi; Locke, Christopher

    2015-07-01

    We study the linear problem associated with modified affine Toda field equation for the Langlands dual gˆ∨, where g ˆ is an untwisted affine Lie algebra. The connection coefficients for the asymptotic solutions of the linear problem are found to correspond to the Q-functions for g-type quantum integrable models. The ψ-system for the solutions associated with the fundamental representations of g leads to Bethe ansatz equations associated with the affine Lie algebra g ˆ . We also study the A2r(2) affine Toda field equation in massless limit in detail and find its Bethe ansatz equations as well as T-Q relations.

  3. Quasi-integrable deformations of the SU(3) Affine Toda theory

    NASA Astrophysics Data System (ADS)

    Ferreira, L. A.; Klimas, P.; Zakrzewski, Wojtek J.

    2016-05-01

    We consider deformations of the SU(3) Affine Toda theory (AT) and investigate the integrability properties of the deformed theories. We find that for some special deformations all conserved quantities change to being conserved only asymptotically, i.e. in the process of the scattering of two solitons these charges do vary in time, but they return, after the scattering, to the values they had prior to the scattering. This phenomenon, which we have called quasi-integrability, is related to special properties of the two-soliton solutions under space-time parity transformations. Some properties of the AT solitons are discussed, especially those involving interesting static multi-soliton solutions. We support our analytical studies with detailed numerical ones in which the time evolution has been simulated by the 4th order Runge-Kutta method. We find that for some perturbations the solitons repel and for the others they form a quasi-bound state. When we send solitons towards each other they can repel when they come close together with or without `flipping' the fields of the model. The solitons radiate very little and appear to be stable. These results support the ideas of quasi-integrability, i.e. that many effects of integrability also approximately hold for the deformed models.

  4. On Affine Fusion and the Phase Model

    NASA Astrophysics Data System (ADS)

    Walton, Mark A.

    2012-11-01

    A brief review is given of the integrable realization of affine fusion discovered recently by Korff and Stroppel. They showed that the affine fusion of the su(n) Wess-Zumino-Novikov-Witten (WZNW) conformal field theories appears in a simple integrable system known as the phase model. The Yang-Baxter equation leads to the construction of commuting operators as Schur polynomials, with noncommuting hopping operators as arguments. The algebraic Bethe ansatz diagonalizes them, revealing a connection to the modular S matrix and fusion of the su(n) WZNW model. The noncommutative Schur polynomials play roles similar to those of the primary field operators in the corresponding WZNW model. In particular, their 3-point functions are the su(n) fusion multiplicities. We show here how the new phase model realization of affine fusion makes obvious the existence of threshold levels, and how it accommodates higher-genus fusion.

  5. Fan Affinity Laws from a Collision Model

    ERIC Educational Resources Information Center

    Bhattacharjee, Shayak

    2012-01-01

    The performance of a fan is usually estimated using hydrodynamical considerations. The calculations are long and involved and the results are expressed in terms of three affinity laws. In this paper we use kinetic theory to attack this problem. A hard sphere collision model is used, and subsequently a correction to account for the flow behaviour…

  6. Intermediate Toda systems

    NASA Astrophysics Data System (ADS)

    Damianou, Pantelis A.; Sabourin, Hervé; Vanhaecke, Pol

    2015-05-01

    We construct a large family of Hamiltonian systems which interpolate between the classical Kostant-Toda lattice and the full Kostant-Toda lattice and we discuss their integrability. There is one such system for every nilpotent ideal I in a Borel subalgebra b+ of an arbitrary simple Lie algebra g. The classical Kostant-Toda lattice corresponds to the case of I = [n+, n+], where n+ is the unipotent ideal of b+, while the full Kostant-Toda lattice corresponds to I = {0}. We mainly focus on the case I = [[n+, n+], n+]. In this case, using the theory of root systems of simple Lie algebras, we compute the rank of the underlying Poisson manifolds and construct a set of (rational) functions in involution, large enough to ensure Liouville integrability. These functions are restrictions of well-chosen integrals of the full Kostant-Toda lattice, except for the case of the Lie algebras of type C and D where a different function (Casimir) is needed. The latter fact, and other ones listed in the paper, point at the Liouville integrability of all the systems we construct, but also at the nontriviality of obtaining the result in full generality.

  7. Flow towards diagonalization for many-body-localization models: adaptation of the Toda matrix differential flow to random quantum spin chains

    NASA Astrophysics Data System (ADS)

    Monthus, Cécile

    2016-07-01

    The iterative methods to diagonalize matrices and many-body Hamiltonians can be reformulated as flows of Hamiltonians towards diagonalization driven by unitary transformations that preserve the spectrum. After a comparative overview of the various types of discrete flows (Jacobi, QR-algorithm) and differential flows (Toda, Wegner, White) that have been introduced in the past, we focus on the random XXZ chain with random fields in order to determine the best closed flow within a given subspace of running Hamiltonians. For the special case of the free-fermion random XX chain with random fields, the flow coincides with the Toda differential flow for tridiagonal matrices which is related to the classical integrable Toda chain and which can be seen as the continuous analog of the discrete QR-algorithm. For the random XXZ chain with random fields that displays a many-body-localization transition, the present differential flow should be an interesting alternative to compare with the discrete flow that has been proposed recently to study the many-body-localization properties in a model of interacting fermions (Rademaker and Ortuno 2016 Phys. Rev. Lett. 116, 010404).

  8. Affine group formulation of the Standard Model coupled to gravity

    SciTech Connect

    Chou, Ching-Yi; Ita, Eyo; Soo, Chopin

    2014-04-15

    In this work we apply the affine group formalism for four dimensional gravity of Lorentzian signature, which is based on Klauder’s affine algebraic program, to the formulation of the Hamiltonian constraint of the interaction of matter and all forces, including gravity with non-vanishing cosmological constant Λ, as an affine Lie algebra. We use the hermitian action of fermions coupled to gravitation and Yang–Mills theory to find the density weight one fermionic super-Hamiltonian constraint. This term, combined with the Yang–Mills and Higgs energy densities, are composed with York’s integrated time functional. The result, when combined with the imaginary part of the Chern–Simons functional Q, forms the affine commutation relation with the volume element V(x). Affine algebraic quantization of gravitation and matter on equal footing implies a fundamental uncertainty relation which is predicated upon a non-vanishing cosmological constant. -- Highlights: •Wheeler–DeWitt equation (WDW) quantized as affine algebra, realizing Klauder’s program. •WDW formulated for interaction of matter and all forces, including gravity, as affine algebra. •WDW features Hermitian generators in spite of fermionic content: Standard Model addressed. •Constructed a family of physical states for the full, coupled theory via affine coherent states. •Fundamental uncertainty relation, predicated on non-vanishing cosmological constant.

  9. Molecular modeling of the affinity chromatography of monoclonal antibodies.

    PubMed

    Paloni, Matteo; Cavallotti, Carlo

    2015-01-01

    Molecular modeling is a methodology that offers the possibility of studying complex systems such as protein-ligand complexes from an atomistic point of view, making available information that can be difficultly obtained from experimental studies. Here, a protocol for the construction of molecular models of the interaction between antibodies and ligands that can be used for an affinity chromatography process is presented. The outlined methodology focuses mostly on the description of a procedure that may be adopted to determine the structure and free energy of interaction between the antibody and the affinity ligand. A procedure to extend the proposed methodology to include the effect of the environment (buffer solution, spacer, support matrix) is also briefly outlined. PMID:25749965

  10. Robust Spectral Clustering Using Statistical Sub-Graph Affinity Model

    PubMed Central

    Eichel, Justin A.; Wong, Alexander; Fieguth, Paul; Clausi, David A.

    2013-01-01

    Spectral clustering methods have been shown to be effective for image segmentation. Unfortunately, the presence of image noise as well as textural characteristics can have a significant negative effect on the segmentation performance. To accommodate for image noise and textural characteristics, this study introduces the concept of sub-graph affinity, where each node in the primary graph is modeled as a sub-graph characterizing the neighborhood surrounding the node. The statistical sub-graph affinity matrix is then constructed based on the statistical relationships between sub-graphs of connected nodes in the primary graph, thus counteracting the uncertainty associated with the image noise and textural characteristics by utilizing more information than traditional spectral clustering methods. Experiments using both synthetic and natural images under various levels of noise contamination demonstrate that the proposed approach can achieve improved segmentation performance when compared to existing spectral clustering methods. PMID:24386111

  11. Local Structural Alignment of RNA with Affine Gap Model

    NASA Astrophysics Data System (ADS)

    Wong, Thomas K. F.; Cheung, Brenda W. Y.; Lam, T. W.; Yiu, S. M.

    Predicting new non-coding RNAs (ncRNAs) of a family can be done by aligning the potential candidate with a member of the family with known sequence and secondary structure. Existing tools either only consider the sequence similarity or cannot handle local alignment with gaps. In this paper, we consider the problem of finding the optimal local structural alignment between a query RNA sequence (with known secondary structure) and a target sequence (with unknown secondary structure) with the affine gap penalty model. We provide the algorithm to solve the problem. Based on a preliminary experiment, we show that there are ncRNA families in which considering local structural alignment with gap penalty model can identify real hits more effectively than using global alignment or local alignment without gap penalty model.

  12. Tending to Change: Toward a Situated Model of Affinity Spaces

    ERIC Educational Resources Information Center

    Bommarito, Dan

    2014-01-01

    The concept of affinity spaces, a theoretical construct used to analyze literate activity from a spatial perspective, has gained popularity among scholars of literacy studies and, particularly, video-game studies. This article seeks to expand current notions of affinity spaces by identifying key assumptions that have limited researchers'…

  13. PREDICTING ER BINDING AFFINITY FOR EDC RANKING AND PRIORITIZATION: MODEL II

    EPA Science Inventory

    The training set used to derive a common reactivity pattern (COREPA) model for estrogen receptor (ER) binding affinity in Model I (see Abstract I in this series) was extended to include 47 rat estrogen receptor (rER) relative binding affinity (RBA) measurements in addition to the...

  14. European bioclimatic affinity groups: Data-model comparisons

    NASA Astrophysics Data System (ADS)

    Laurent, J.-M.; François, L.; Bar-Hen, A.; Bel, L.; Cheddadi, R.

    2008-03-01

    Global vegetation models are remarkably effective when considering large areas such as Europe. However, their accuracy at finer scales remains to be tested. In this paper, we validate the simulation of modern potential vegetation by the CARbon Assimilation In the Biosphere (CARAIB) model in Europe. Then, in order to evaluate the simulation of tree group distributions at a finer scale, in France, we present a comparison between observed distributions, distributions reconstructed from palynological data, and model simulated ranges. The results will help to validate past vegetation simulations. For this analysis, we use Bioclimatic Affinity Groups (BAGs), based on vegetation groups' climatic tolerances and requirements. The CARAIB model was adapted to simulate the net primary productivity (NPP), biomass and range of the arboreal BAGs. In Europe, at a 30' latitude/longitude grid scale, simulated NPP of BAGs are used to define classes of vegetation as being present or absent, with a classification rule, based on Kappa statistics. In France, at a 10' lat./long. scale, a second discriminant analysis, based on Classification And Regression Tree (CART), allows for a similar classification with BAG pollen percentages. At each palynological sampling site, we then compared the simulation to the reconstruction from pollen data. With 30' lat./long. resolution, most thresholds that discriminate NPP into absence or presence classes are low, ranging from 1 to 77 g/m 2. Agreement indices between observed and simulated distributions range from 0.4 to 0.83, with broad scale BAG potential patterns and boundaries being accurately simulated by CARAIB. In France, on the 10' lat./long. scale, pollen percentages correctly account for BAG presence/absence despite non-linear pollen-vegetation relationships. Agreement ratios between observed and reconstructed patterns range from 0.53 to 0.95. At the 10' lat./long. scale, the validation of simulated ranges with pollen data is reliable for 9 of

  15. Mathematical model accurately predicts protein release from an affinity-based delivery system.

    PubMed

    Vulic, Katarina; Pakulska, Malgosia M; Sonthalia, Rohit; Ramachandran, Arun; Shoichet, Molly S

    2015-01-10

    Affinity-based controlled release modulates the delivery of protein or small molecule therapeutics through transient dissociation/association. To understand which parameters can be used to tune release, we used a mathematical model based on simple binding kinetics. A comprehensive asymptotic analysis revealed three characteristic regimes for therapeutic release from affinity-based systems. These regimes can be controlled by diffusion or unbinding kinetics, and can exhibit release over either a single stage or two stages. This analysis fundamentally changes the way we think of controlling release from affinity-based systems and thereby explains some of the discrepancies in the literature on which parameters influence affinity-based release. The rate of protein release from affinity-based systems is determined by the balance of diffusion of the therapeutic agent through the hydrogel and the dissociation kinetics of the affinity pair. Equations for tuning protein release rate by altering the strength (KD) of the affinity interaction, the concentration of binding ligand in the system, the rate of dissociation (koff) of the complex, and the hydrogel size and geometry, are provided. We validated our model by collapsing the model simulations and the experimental data from a recently described affinity release system, to a single master curve. Importantly, this mathematical analysis can be applied to any single species affinity-based system to determine the parameters required for a desired release profile. PMID:25449806

  16. Kp and Toda Tau Functions in Bethe Ansatz

    NASA Astrophysics Data System (ADS)

    Takasaki, Kanehisa

    2011-10-01

    Recent work of Foda and his group on a connection between classical integrable hierarchies (the KP and 2D Toda hierarchies) and some quantum integrable systems (the 6-vertex model with DWBC, the finite XXZ chain of spin 1/2, the phase model on a finite chain, etc.) is reviewed. Some additional information on this issue is also presented.

  17. Disorder, pre-stress and non-affinity in polymer 8-chain models

    NASA Astrophysics Data System (ADS)

    Cioroianu, Adrian R.; Spiesz, Ewa M.; Storm, Cornelis

    2016-04-01

    To assess the role of single-chain elasticity, non-affine strain fields and pre-stressed reference states we present and discuss the results of numerical and analytical analyses of a modified 8-chain Arruda-Boyce model for cross-linked polymer networks. This class of models has proved highly successful in modeling the finite-strain response of flexible rubbers. We extend it to include the effects of spatial disorder and the associated non-affinity, and use it to assess the validity of replacing the constituent chain's nonlinear elastic response with equivalent linear, Hookean springs. Surprisingly, we find that even in the regime of linear response, the full polymer model gives very different results from its linearized counterpart, even though none of the chains are stretched beyond their linear regime. We demonstrate that this effect is due to the fact that the polymer models are under considerable pre-stress in their ground state. We show that pre-stress strongly suppresses non-affinity in these unit cell models, resulting in a marked stiffening of the bulk response. Polymer networks with some degree of flexibility are thus intrinsically prestressed, and one effect of such prestresses is to reduce non-affine deformations. Combined, these findings may help explain why fully affine mechanical models, in many cases, predict the bulk mechanical response of disordered polymer networks so well.

  18. Toda Systems, Cluster Characters, and Spectral Networks

    NASA Astrophysics Data System (ADS)

    Williams, Harold

    2016-07-01

    We show that the Hamiltonians of the open relativistic Toda system are elements of the generic basis of a cluster algebra, and in particular are cluster characters of nonrigid representations of a quiver with potential. Using cluster coordinates defined via spectral networks, we identify the phase space of this system with the wild character variety related to the periodic nonrelativistic Toda system by the wild nonabelian Hodge correspondence. We show that this identification takes the relativistic Toda Hamiltonians to traces of holonomies around a simple closed curve. In particular, this provides nontrivial examples of cluster coordinates on SL n -character varieties for n > 2 where canonical functions associated to simple closed curves can be computed in terms of quivers with potential, extending known results in the SL 2 case.

  19. Determination of protein-ligand binding affinity by NMR: observations from serum albumin model systems.

    PubMed

    Fielding, Lee; Rutherford, Samantha; Fletcher, Dan

    2005-06-01

    The usefulness of bovine serum albumin (BSA) as a model protein for testing NMR methods for the study of protein-ligand interactions is discussed. Isothermal titration calorimetry established the binding affinity and stoichiometry of the specific binding site for L-tryptophan, D-tryptophan, naproxen, ibuprofen, salicylic acid and warfarin. The binding affinities of the same ligands determined by NMR methods are universally weaker (larger KD). This is because the NMR methods are susceptible to interference from additional non-specific binding. The L-tryptophan-BSA and naproxen-BSA systems were the best behaved model systems. PMID:15816062

  20. Predicting binding affinities of diverse pharmaceutical chemicals to human serum plasma proteins using QSPR modelling approaches.

    PubMed

    Basant, N; Gupta, S; Singh, K P

    2016-01-01

    The prediction of the plasma protein binding (PPB) affinity of chemicals is of paramount significance in the drug development process. In this study, ensemble machine learning-based QSPR models have been established for a four-category classification and PPB affinity prediction of diverse compounds using a large PPB dataset of 930 compounds and in accordance with the OECD guidelines. The structural diversity of the chemicals was tested by the Tanimoto similarity index. The external predictive power of the developed QSPR models was evaluated through internal and external validations. In the QSPR models, XLogP was the most important descriptor. In the test data, the classification QSPR models rendered an accuracy of >93%, while the regression QSPR models yielded r(2) of >0.920 between the measured and predicted PPB affinities, with the root mean squared error <9.77. Values of statistical coefficients derived for the test data were above their threshold limits, thus put a high confidence in this analysis. The QSPR models in this study performed better than any of the previous studies. The results suggest that the developed QSPR models are reliable for predicting the PPB affinity of structurally diverse chemicals. They can be useful for initial screening of candidate molecules in the drug development process. PMID:26854728

  1. The extended Z N -Toda hierarchy

    NASA Astrophysics Data System (ADS)

    Li, Chuanzhong; He, Jingsong

    2015-11-01

    We construct the extended flow equations of a new Z N -Toda hierarchy taking values in a commutative subalgebra Z N of gl( N, C). We give the Hirota bilinear equations and tau function of this new extended Z N -Toda hierarchy. Taking the presence of logarithmic terms into account, we construct some extended vertex operators in generalized Hirota bilinear equations, which might be useful in topological field theory and the Gromov-Witten theory. We present the Darboux transformations and bi-Hamiltonian structure of this hierarchy. Using Hamiltonian tau-symmetry, we obtain another tau function of this hierarchy with some unknown mysterious relation to the tau function derived using the Sato theory.

  2. Generalised Eisenhart lift of the Toda chain

    SciTech Connect

    Cariglia, Marco; Gibbons, Gary

    2014-02-15

    The Toda chain of nearest neighbour interacting particles on a line can be described both in terms of geodesic motion on a manifold with one extra dimension, the Eisenhart lift, or in terms of geodesic motion in a symmetric space with several extra dimensions. We examine the relationship between these two realisations and discover that the symmetric space is a generalised, multi-particle Eisenhart lift of the original problem that reduces to the standard Eisenhart lift. Such generalised Eisenhart lift acts as an inverse Kaluza-Klein reduction, promoting coupling constants to momenta in higher dimension. In particular, isometries of the generalised lift metric correspond to energy preserving transformations that mix coordinates and coupling constants. A by-product of the analysis is that the lift of the Toda Lax pair can be used to construct higher rank Killing tensors for both the standard and generalised lift metrics.

  3. A diffusion model for picosecond electron bunches from negative electron affinity GaAs photo cathodes

    SciTech Connect

    P. Hartmann; J. Bermuth; D. v. Harrach; J. Hoffmann; S. Kobis; E. Reichert; K. Aulenbacher; J. Schuler; M. Steigerwald

    1998-10-27

    Even though theoretical estimates predict response times for the photo emission process of electrons from a negative electron affinity GaAs photo emitter in excess of hundreds of picoseconds, recent measurements found electron bunch durations of 40 ps or less. This work presents precise measurements of picosecond electron bunches from a negative affinity bulk GaAs photo cathode and develops a model which explains the measured bunch durations as well as the observed bunch shapes. The bunch shape turns out to be independent from the quantum efficiency of the photo emitter.

  4. Singular structure of Toda lattices and cohomology of certain compact Lie groups

    NASA Astrophysics Data System (ADS)

    Casian, Luis; Kodama, Yuji

    2007-05-01

    We study the singularities (blow-ups) of the Toda lattice associated with a real split semisimple Lie algebra . It turns out that the total number of blow-up points along trajectories of the Toda lattice is given by the number of points of a Chevalley group related to the maximal compact subgroup K of the group with over the finite field . Here is the Langlands dual of E The blow-ups of the Toda lattice are given by the zero set of the [tau]-functions. For example, the blow-ups of the Toda lattice of A-type are determined by the zeros of the Schur polynomials associated with rectangular Young diagrams. Those Schur polynomials are the [tau]-functions for the nilpotent Toda lattices. Then we conjecture that the number of blow-ups is also given by the number of real roots of those Schur polynomials for a specific variable. We also discuss the case of periodic Toda lattice in connection with the real cohomology of the flag manifold associated to an affine Kac-Moody algebra.

  5. Performance comparison of rigid and affine models for motion estimation using ultrasound radio-frequency signals.

    PubMed

    Pan, Xiaochang; Liu, Ke; Shao, Jinghua; Gao, Jing; Huang, Lingyun; Bai, Jing; Luo, Jianwen

    2015-11-01

    Tissue motion estimation is widely used in many ultrasound techniques. Rigid-model-based and nonrigid-modelbased methods are two main groups of space-domain methods of tissue motion estimation. The affine model is one of the commonly used nonrigid models. The performances of the rigid model and affine model have not been compared on ultrasound RF signals, which have been demonstrated to obtain higher accuracy, precision, and resolution in motion estimation compared with B-mode images. In this study, three methods, i.e., the normalized cross-correlation method with rigid model (NCC), the optical flow method with rigid model (OFRM), and the optical flow method with affine model (OFAM), are compared using ultrasound RF signals, rather than the B-mode images used in previous studies. Simulations, phantom, and in vivo experiments are conducted to make the comparison. In the simulations, the root-mean-square errors (RMSEs) of axial and lateral displacements and strains are used to assess the accuracy of motion estimation, and the elastographic signal-tonoise ratio (SNRe) and contrast-to-noise ratio (CNRe) are used to evaluate the quality of axial strain images. In the phantom experiments, the registration error between the pre- and postdeformation RF signals, as well as the SNRe and CNRe of axial strain images, are utilized as the evaluation criteria. In the in vivo experiments, the registration error is used to evaluate the estimation performance. The results show that the affinemodel- based method (i.e., OFAM) obtains the lowest RMSE or registration error and the highest SNRe and CNRe among all the methods. The affine model is demonstrated to be superior to the rigid model in motion estimation based on RF signals. PMID:26559623

  6. Smooth bounce in the affine quantization of a Bianchi I model

    NASA Astrophysics Data System (ADS)

    Bergeron, Hervé; Dapor, Andrea; Gazeau, Jean Pierre; Małkiewicz, Przemysław

    2015-06-01

    We present the affine coherent state quantization of the Bianchi I model. As in our previous paper on quantum theory of Friedmann models, we employ a variable associated with a perfect fluid to play a role of clock. Then we deparametrize the model. A distinctive feature, absent in isotropic models, is an extra nonholonomic constraint, which survives the deparametrization and constrains the range of physical variables. The appearance of the constraint reflects the "amplification" of singularity due to anisotropy. The quantization smoothes the extra constraint and allows quantum contracting trajectories to be smoothly transformed into expanding ones. Making use of an affine coherent state we develop a semiclassical description. Figures are included to illustrate our result.

  7. Self-affine and ARX-models zonation of well logging data

    NASA Astrophysics Data System (ADS)

    Shiri, Yousef; Tokhmechi, Behzad; Zarei, Zeinab; Koneshloo, Mohammad

    2012-11-01

    Zonation of time series into models which their parameters are piecewise constant are important and well-studied problems. Geophysical well logging data often show a complex pattern due to their multifractal nature. In a multifractal system, any pieces of it are established by a distinct exponent that can characterize them. This feature has the capability to cluster them. Self-affine zonation by Auto Regressive model with exogenous inputs (ARX) is a new approach which places well logging segments in the clusters which are more self-affine against the other clusters. This approach was performed and compared with a conventional ARX zonation in the well logging data of three different oilfields in southern parts of Iran. The results showed a good accuracy for detecting homogeneous lithological segments and led to a precise interpretation process to update the reservoir architecture.

  8. Maximum-Entropy Models of Sequenced Immune Repertoires Predict Antigen-Antibody Affinity.

    PubMed

    Asti, Lorenzo; Uguzzoni, Guido; Marcatili, Paolo; Pagnani, Andrea

    2016-04-01

    The immune system has developed a number of distinct complex mechanisms to shape and control the antibody repertoire. One of these mechanisms, the affinity maturation process, works in an evolutionary-like fashion: after binding to a foreign molecule, the antibody-producing B-cells exhibit a high-frequency mutation rate in the genome region that codes for the antibody active site. Eventually, cells that produce antibodies with higher affinity for their cognate antigen are selected and clonally expanded. Here, we propose a new statistical approach based on maximum entropy modeling in which a scoring function related to the binding affinity of antibodies against a specific antigen is inferred from a sample of sequences of the immune repertoire of an individual. We use our inference strategy to infer a statistical model on a data set obtained by sequencing a fairly large portion of the immune repertoire of an HIV-1 infected patient. The Pearson correlation coefficient between our scoring function and the IC50 neutralization titer measured on 30 different antibodies of known sequence is as high as 0.77 (p-value 10-6), outperforming other sequence- and structure-based models. PMID:27074145

  9. Maximum-Entropy Models of Sequenced Immune Repertoires Predict Antigen-Antibody Affinity

    PubMed Central

    Marcatili, Paolo; Pagnani, Andrea

    2016-01-01

    The immune system has developed a number of distinct complex mechanisms to shape and control the antibody repertoire. One of these mechanisms, the affinity maturation process, works in an evolutionary-like fashion: after binding to a foreign molecule, the antibody-producing B-cells exhibit a high-frequency mutation rate in the genome region that codes for the antibody active site. Eventually, cells that produce antibodies with higher affinity for their cognate antigen are selected and clonally expanded. Here, we propose a new statistical approach based on maximum entropy modeling in which a scoring function related to the binding affinity of antibodies against a specific antigen is inferred from a sample of sequences of the immune repertoire of an individual. We use our inference strategy to infer a statistical model on a data set obtained by sequencing a fairly large portion of the immune repertoire of an HIV-1 infected patient. The Pearson correlation coefficient between our scoring function and the IC50 neutralization titer measured on 30 different antibodies of known sequence is as high as 0.77 (p-value 10−6), outperforming other sequence- and structure-based models. PMID:27074145

  10. A deformation of quantum affine algebra in squashed Wess-Zumino-Novikov-Witten models

    SciTech Connect

    Kawaguchi, Io; Yoshida, Kentaroh

    2014-06-01

    We proceed to study infinite-dimensional symmetries in two-dimensional squashed Wess-Zumino-Novikov-Witten models at the classical level. The target space is given by squashed S³ and the isometry is SU(2){sub L}×U(1){sub R}. It is known that SU(2){sub L} is enhanced to a couple of Yangians. We reveal here that an infinite-dimensional extension of U(1){sub R} is a deformation of quantum affine algebra, where a new deformation parameter is provided with the coefficient of the Wess-Zumino term. Then we consider the relation between the deformed quantum affine algebra and the pair of Yangians from the viewpoint of the left-right duality of monodromy matrices. The integrable structure is also discussed by computing the r/s-matrices that satisfy the extended classical Yang-Baxter equation. Finally, two degenerate limits are discussed.

  11. Probabilistic approach for predicting periodic orbits in piecewise affine differential models.

    PubMed

    Chaves, Madalena; Farcot, Etienne; Gouzé, Jean-Luc

    2013-06-01

    Piecewise affine models provide a qualitative description of the dynamics of a system, and are often used to study genetic regulatory networks. The state space of a piecewise affine system is partitioned into hyperrectangles, which can be represented as nodes in a directed graph, so that the system's trajectories follow a path in a transition graph. This paper proposes and compares two definitions of probability of transition between two nodes A and B of the graph, based on the volume of the initial conditions on the hyperrectangle A whose trajectories cross to B. The parameters of the system can thus be compared to the observed transitions between two hyperrectangles. This property may become useful to identify sets of parameters for which the system yields a desired periodic orbit with a high probability, or to predict the most likely periodic orbit given a set of parameters, as illustrated by a gene regulatory system composed of two intertwined negative loops. PMID:23054666

  12. Modelling the binding affinity of steroids to zebrafish sex hormone-binding globulin.

    PubMed

    Saxena, A K; Devillers, J; Pery, A R R; Beaudouin, R; Balaramnavar, V M; Ahmed, S

    2014-01-01

    The circulating endogenous steroids are transported in the bloodstream. These are bound to a highly specific sex hormone-binding globulin (SHBG) and in lower affinity to proteins such as the corticosteroid-binding protein and albumin in vertebrates, including fish. It is generally believed that the glycoprotein SHBG protects these steroids from rapid metabolic degradation and thus intervenes in its availability at the target tissues. Endocrine disrupters binding to SHBG affect the normal activity of natural steroids. Since xenobiotics are primarily released in the aquatic environment, there is a need to evaluate the binding affinity of xenosteroid mimics on fish SHBG, especially in zebrafish (Danio rerio), a small freshwater fish originating in India and widely employed in ecotoxicology, toxicology, and genetics. In this context, a zebrafish SHBG (zfSHBG) homology model was developed using the human SHBG (hSHBG) receptor structure as template. It was shown that interactions with amino acids Ser-36, Asp-59 and Thr-54 were important for binding affinity. A ligand-based pharmacophore model was also developed for both zfSHBG and hSHBG inhibitors that differentiated binders from non-binders, but also demonstrated structural requirements for zfSHBG and hSHBG ligands. The study provides insights into the mechanism of action of endocrine disruptors in zebrafish as well as providing a useful tool for identifying anthropogenic compounds inhibiting zfSHBG. PMID:24874994

  13. Microarrays as Model Biosensor Platforms to Investigate the Structure and Affinity of Aptamers.

    PubMed

    Martin, Jennifer A; Chushak, Yaroslav; Chávez, Jorge L; Hagen, Joshua A; Kelley-Loughnane, Nancy

    2016-01-01

    Immobilization of nucleic acid aptamer recognition elements selected free in solution onto the surface of biosensor platforms has proven challenging. This study investigated the binding of multiple aptamer/target pairs immobilized on a commercially available microarray as a model system mimicking biosensor applications. The results indicate a minimum distance (linker length) from the surface and thymine nucleobase linker provides reproducible binding across varying conditions. An indirect labeling method, where the target was labeled with a biotin followed by a brief Cy3-streptavidin incubation, provided a higher signal-to-noise ratio and over two orders of magnitude improvement in limit of detection, compared to direct Cy3-protein labeling. We also showed that the affinities of the aptamer/target interaction can change between direct and indirect labeling and conditions to optimize for the highest fluorescence intensity will increase the sensitivity of the assay but will not change the overall affinity. Additionally, some sequences which did not initially bind demonstrated binding when conditions were optimized. These results, in combination with studies demonstrating enhanced binding in nonselection buffers, provided insights into the structure and affinity of aptamers critical for biosensor applications and allowed for generalizations in starting conditions for researchers wishing to investigate aptamers on a microarray surface. PMID:27042344

  14. Microarrays as Model Biosensor Platforms to Investigate the Structure and Affinity of Aptamers

    PubMed Central

    Martin, Jennifer A.; Chushak, Yaroslav; Chávez, Jorge L.; Hagen, Joshua A.; Kelley-Loughnane, Nancy

    2016-01-01

    Immobilization of nucleic acid aptamer recognition elements selected free in solution onto the surface of biosensor platforms has proven challenging. This study investigated the binding of multiple aptamer/target pairs immobilized on a commercially available microarray as a model system mimicking biosensor applications. The results indicate a minimum distance (linker length) from the surface and thymine nucleobase linker provides reproducible binding across varying conditions. An indirect labeling method, where the target was labeled with a biotin followed by a brief Cy3-streptavidin incubation, provided a higher signal-to-noise ratio and over two orders of magnitude improvement in limit of detection, compared to direct Cy3-protein labeling. We also showed that the affinities of the aptamer/target interaction can change between direct and indirect labeling and conditions to optimize for the highest fluorescence intensity will increase the sensitivity of the assay but will not change the overall affinity. Additionally, some sequences which did not initially bind demonstrated binding when conditions were optimized. These results, in combination with studies demonstrating enhanced binding in nonselection buffers, provided insights into the structure and affinity of aptamers critical for biosensor applications and allowed for generalizations in starting conditions for researchers wishing to investigate aptamers on a microarray surface. PMID:27042344

  15. On the computation of stress in affine versus nonaffine fibril kinematics within planar collagen network models.

    PubMed

    Pence, Thomas J; Monroe, Ryan J; Wright, Neil T

    2008-08-01

    Some recent analyses modeled the response of collagenous tissues, such as epicardium, using a hypothetical network consisting of interconnected springlike fibers. The fibers in the network were organized such that internal nodes served as the connection point between three such collagen springs. The results for assumed affine and nonaffine deformations are contrasted after a homogeneous deformation at the boundary. Affine deformation provides a stiffer mechanical response than nonaffine deformation. In contrast to nonaffine deformation, affine deformation determines the displacement of internal nodes without imposing detailed force balance, thereby complicating the simplest intuitive notion of stress, one based on free body cuts, at the single node scale. The standard notion of stress may then be recovered via average field theory computations based on large micromesh realizations. An alternative and by all indications complementary viewpoint for the determination of stress in these collagen fiber networks is discussed here, one in which stress is defined using elastic energy storage, a notion which is intuitive at the single node scale. It replaces the average field theory computations by an averaging technique over randomly oriented isolated simple elements. The analytical operations do not require large micromesh realizations, but the tedious nature of the mathematical manipulation is clearly aided by symbolic algebra calculation. For the example case of linear elastic deformation, this results in material stiffnesses that relate the infinitesimal strain and stress. The result that the affine case is stiffer than the nonaffine case is recovered, as would be expected. The energy framework also lends itself to the natural inclusion of changes in mechanical response due to the chemical, electrical, or thermal environment. PMID:18601451

  16. From Toda to KdV

    NASA Astrophysics Data System (ADS)

    Bambusi, D.; Kappeler, T.; Paul, T.

    2015-07-01

    For periodic Toda chains with a large number N of particles we consider states which are N-2-close to the equilibrium and constructed by discretizing arbitrary given C2-functions with mesh size N-1. Our aim is to describe the spectrum of the Jacobi matrices LN appearing in the Lax pair formulation of the dynamics of these states as N → ∞. To this end we construct two Hill operators H±—such operators come up in the Lax pair formulation of the Korteweg-de Vries equation—and prove by methods of semiclassical analysis that the asymptotics as N → ∞ of the eigenvalues at the edges of the spectrum of LN are of the form +/- (2-(2N)-2 λ ^+/- n + \\cdots ) where (λ ^+/- _n)n ≥ 0 are the eigenvalues of H±. In the bulk of the spectrum, the eigenvalues are o(N-2)-close to the ones of the equilibrium matrix. As an application we obtain asymptotics of a similar type of the discriminant, associated to LN.

  17. Nonlinear force propagation, anisotropic stiffening and non-affine relaxation in a model cytoskeleton

    NASA Astrophysics Data System (ADS)

    Mizuno, Daisuke; Head, David; Ikebe, Emi; Nakamasu, Akiko; Kinoshita, Suguru; Peijuan, Zhang; Ando, Shoji

    2013-03-01

    Forces are generated heterogeneously in living cells and transmitted through cytoskeletal networks that respond highly non-linearly. Here, we carry out high-bandwidth passive microrheology on vimentin networks reconstituted in vitro, and observe the nonlinear mechanical response due to forces propagating from a local source applied by an optical tweezer. Since the applied force is constant, the gel becomes equilibrated and the fluctuation-dissipation theorem can be employed to deduce the viscoelasticity of the local environment from the thermal fluctuations of colloidal probes. Our experiments unequivocally demonstrate the anisotropic stiffening of the cytoskeletal network behind the applied force, with greater stiffening in the parallel direction. Quantitative agreement with an affine continuum model is obtained, but only for the response at certain frequency ~ 10-1000 Hz which separates the high-frequency power law and low-frequency elastic behavior of the network. We argue that the failure of the model at lower frequencies is due to the presence of non-affinity, and observe that zero-frequency changes in particle separation can be fitted when an independently-measured, empirical nonaffinity factor is applied.

  18. Copper binding to soil fulvic and humic acids: NICA-Donnan modeling and conditional affinity spectra.

    PubMed

    Xu, Jinling; Tan, Wenfeng; Xiong, Juan; Wang, Mingxia; Fang, Linchuan; Koopal, Luuk K

    2016-07-01

    Binding of Cu(II) to soil fulvic acid (JGFA), soil humic acids (JGHA, JLHA), and lignite-based humic acid (PAHA) was investigated through NICA-Donnan modeling and conditional affinity spectrum (CAS). It is to extend the knowledge of copper binding by soil humic substances (HS) both in respect of enlarging the database of metal ion binding to HS and obtaining a good insight into Cu binding to the functional groups of FA and HA by using the NICA-Donnan model to unravel the intrinsic and conditional affinity spectra. Results showed that Cu binding to HS increased with increasing pH and decreasing ionic strength. The amount of Cu bound to the HAs was larger than the amount bound to JGFA. Milne's generic parameters did not provide satisfactory predictions for the present soil HS samples, while material-specific NICA-Donnan model parameters described and predicted Cu binding to the HS well. Both the 'low' and 'high' concentration fitting procedures indicated a substantial bidentate structure of the Cu complexes with HS. By means of CAS underlying NICA isotherm, which was scarcely used, the nature of the binding at different solution conditions for a given sample and the differences in binding mode were illustrated. It was indicated that carboxylic group played an indispensable role in Cu binding to HS in that the carboxylic CAS had stronger conditional affinity than the phenolic distribution due to its large degree of proton dissociation. The fact was especially true for JGFA and JLHA which contain much larger amount of carboxylic groups, and the occupation of phenolic sites by Cu was negligible. Comparable amounts of carboxylic and phenolic groups on PAHA and JGHA, increased the occupation of phenolic type sites by Cu. The binding strength of PAHA-Cu and JGHA-Cu was stronger than that of JGFA-Cu and JLHA-Cu. The presence of phenolic groups increased the chance of forming more stable complexes, such as the salicylate-Cu or catechol-Cu type structures. PMID:27061366

  19. Affinity comparison of different THCA synthase to CBGA using modeling computational approaches

    PubMed Central

    Alaoui, Moulay Abdelaziz El; Ibrahimi, Azeddine; Semlali, Oussama; Tarhda, Zineb; Marouane, Melloul; Najwa, Alaoui; Soulaymani, Abdelmajid; Fahime, Elmostafa El

    2014-01-01

    The Δ9-Tetrahydrocannabinol (THCA) is the primary psychoactive compound of Cannabis Sativa. It is produced by Δ1- Tetrahydrocannabinolic acid synthase (THCA) which catalyzes the oxidative cyclization of cannabigerolic acid (CBGA) the precursor of the THCA. In this study, we were interested by the three dimensional structure of THCA synthase protein. Generation of models were done by MODELLER v9.11 and homology modeling with Δ1-tetrahydrocannabinolic acid (THCA) synthase X ray structure (PDB code 3VTE) on the basis of sequences retrieved from GenBank. Procheck, Errat, and Verify 3D tools were used to verify the reliability of the six 3D models obtained, the overall quality factor and the Prosa Z-score were also used to check the quality of the six modeled proteins. The RMSDs for C-alpha atoms, main-chain atoms, side-chain atoms and all atoms between the modeled structures and the corresponding template ranged between 0.290 Å-1.252 Å, reflecting the good quality of the obtained models. Our study of the CBGA-THCA synthase docking demonstrated that the active site pocket was successfully recognized using computational approach. The interaction energy of CBGA computed in ‘fiber types’ proteins ranged between -4.1 95 kcal/mol and -5.95 kcal/mol whereas in the ‘drug type’ was about -7.02 kcal/mol to -7.16 kcal/mol, which maybe indicate the important role played by the interaction energy of CBGA in the determination of the THCA level in Cannabis Sativa L. varieties. Finally, we have proposed an experimental design in order to explore the binding energy source of ligand-enzyme in Cannabis Sativa and the production level of the THCA in the absence of any information regarding the correlation between the enzyme affinity and THCA level production. This report opens the doors to more studies predicting the binding site pocket with accuracy from the perspective of the protein affinity and THCA level produced in Cannabis Sativa. PMID:24516324

  20. Affinity Chromatography.

    ERIC Educational Resources Information Center

    Gray, Gary R.

    1980-01-01

    Presents selected recent advances in immobilization chemistry which have important connections to affinity chromatography. Discusses ligand immobilization and support modification. Cites 51 references. (CS)

  1. A Molecular Mechanics Approach to Modeling Protein-Ligand Interactions: Relative Binding Affinities in Congeneric Series

    PubMed Central

    Rapp, Chaya S.; Kalyanaraman, Chakrapani; Schiffmiller, Aviva; Schoenbrun, Esther Leah; Jacobson, Matthew P.

    2011-01-01

    We introduce the “Prime-ligand” method for ranking ligands in congeneric series. The method employs a single scoring function, the OPLS-AA/GBSA molecular mechanics/implicit solvent model, for all stages of sampling and scoring. We evaluate the method using 12 test sets of congeneric series for which experimental binding data is available in the literature, as well as the structure of one member of the series bound to the protein. Ligands are ‘docked’ by superimposing a common stem fragment among the compounds in the series using a crystal complex from the Protein Databank, and sampling the conformational space of the variable region. Our results show good correlation between our predicted rankings and experimental data for cases in which binding affinities differ by at least one order of magnitude. For 11 out of 12 cases, >90% of such ligand pairs could be correctly ranked, while for the remaining case, Factor Xa, 76% of such pairs were correctly ranked. A small number of compounds could not be docked using the current protocol due to the large size of functional groups that could not be accommodated by a rigid receptor. CPU requirements for the method, involving CPU-minutes per ligand, are modest compared with more rigorous methods that use similar force fields, such as free energy perturbation. We also benchmark the scoring function using series of ligand bound to the same protein within the CSAR data set. We demonstrate that energy minimization of ligand in the crystal structures is critical to obtain any correlation with experimentally determined binding affinities. PMID:21780805

  2. Cluster characters and the combinatorics of Toda systems

    NASA Astrophysics Data System (ADS)

    Williams, H.

    2015-12-01

    We survey some connections between Toda systems and cluster algebras. One of these connections is based on representation theory: it is known that Laurent expansions of cluster variables are generating functions of Euler characteristics of quiver Grassmannians, and the same turns out to be true of the Hamiltonians of the open relativistic Toda chain. Another connection is geometric: the closed nonrelativistic Toda chain can be regarded as a meromorphic Hitchin system and studied from the standpoint of spectral networks. From this standpoint, the combinatorial formulas for the Hamiltonians of the open relativistic system are sums of trajectories of differential equations defined by the closed nonrelativistic spectral curves.

  3. Spatially Enhanced Differential RNA Methylation Analysis from Affinity-Based Sequencing Data with Hidden Markov Model

    PubMed Central

    Zhang, Yu-Chen; Zhang, Shao-Wu; Liu, Lian; Liu, Hui; Zhang, Lin; Cui, Xiaodong; Huang, Yufei; Meng, Jia

    2015-01-01

    With the development of new sequencing technology, the entire N6-methyl-adenosine (m6A) RNA methylome can now be unbiased profiled with methylated RNA immune-precipitation sequencing technique (MeRIP-Seq), making it possible to detect differential methylation states of RNA between two conditions, for example, between normal and cancerous tissue. However, as an affinity-based method, MeRIP-Seq has yet provided base-pair resolution; that is, a single methylation site determined from MeRIP-Seq data can in practice contain multiple RNA methylation residuals, some of which can be regulated by different enzymes and thus differentially methylated between two conditions. Since existing peak-based methods could not effectively differentiate multiple methylation residuals located within a single methylation site, we propose a hidden Markov model (HMM) based approach to address this issue. Specifically, the detected RNA methylation site is further divided into multiple adjacent small bins and then scanned with higher resolution using a hidden Markov model to model the dependency between spatially adjacent bins for improved accuracy. We tested the proposed algorithm on both simulated data and real data. Result suggests that the proposed algorithm clearly outperforms existing peak-based approach on simulated systems and detects differential methylation regions with higher statistical significance on real dataset. PMID:26301253

  4. 2d Affine XY-Spin Model/4d Gauge Theory Duality and Deconfinement

    SciTech Connect

    Anber, Mohamed M.; Poppitz, Erich; Unsal, Mithat; /SLAC /Stanford U., Phys. Dept. /San Francisco State U.

    2012-08-16

    We introduce a duality between two-dimensional XY-spin models with symmetry-breaking perturbations and certain four-dimensional SU(2) and SU(2) = Z{sub 2} gauge theories, compactified on a small spatial circle R{sup 1,2} x S{sup 1}, and considered at temperatures near the deconfinement transition. In a Euclidean set up, the theory is defined on R{sup 2} x T{sup 2}. Similarly, thermal gauge theories of higher rank are dual to new families of 'affine' XY-spin models with perturbations. For rank two, these are related to models used to describe the melting of a 2d crystal with a triangular lattice. The connection is made through a multi-component electric-magnetic Coulomb gas representation for both systems. Perturbations in the spin system map to topological defects in the gauge theory, such as monopole-instantons or magnetic bions, and the vortices in the spin system map to the electrically charged W-bosons in field theory (or vice versa, depending on the duality frame). The duality permits one to use the two-dimensional technology of spin systems to study the thermal deconfinement and discrete chiral transitions in four-dimensional SU(N{sub c}) gauge theories with n{sub f} {ge} 1 adjoint Weyl fermions.

  5. Development of novel cellular model for affinity studies of histamine H(4) receptor ligands.

    PubMed

    Karcz, Tadeusz; Kieć-Kononowicz, Katarzyna

    2013-01-01

    The G protein-coupled histamine H4 receptor (H4R) is the last member of histamine receptors family discovered so far. Its expression pattern, together with postulated involvement in a wide variety of immunological and inflammatory processes make histamine H4 receptor an interesting target for drug development. Potential H4R ligands may provide an innovative therapies for different immuno-based diseases, including allergy, asthma, pruritus associated with allergy or autoimmune skin conditions, rheumatoid arthritis and pain. However, none of successfully developed selective and potent histamine H4 receptor ligands have been introduced to the market up to date. For that reason there is still a strong demand for pharmacological models to be used in studies on potent H4R ligands. In current work we present the development of novel mammalian cell line, stably expressing human histamine H4 receptor, with use of retroviral transduction approach. Obtained cell line was pharmacologically characterized in radioligand binding studies and its utility for affinity testing of potent receptor ligands was confirmed in comparative studies with the use of relevant insect cells expression model. Obtained results allow for statement that developed cellular model may be successfully employed in search for new compounds active at histamine H4 receptor. PMID:24432340

  6. Orthogonal polynomial interpretation of Δ-Toda equations

    NASA Astrophysics Data System (ADS)

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

    2015-10-01

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

  7. Tumor affinity of radiolabeled peanut agglutinin compared with that of Ga-67 citrate in animal models

    SciTech Connect

    Yokoyama, K.; Aburano, T.; Watanabe, N.; Kawabata, S.; Ishida, H.; Mukai, K.; Tonami, N.; Hisada, K.

    1985-05-01

    Peanut agglutinin (PNA) binds avidly to the immunodominant group of the tumor associated T antigen. The purpose of this study was to evaluate oncodiagnostic potential of radiolabeled PNA in animal models. PNA was labeled with I-125 or I-131 by Iodogen and also with In-111 by cyclic DTPA anhydride. The biological activity of PNA was examined by a hemaglutination titer with a photometer before and after labeling. Animal tumor models used were Lewis Lung Cancer(LLC), B-16 Melanotic Melanoma(MM), Yoshida Sarcoma(YS), Ehrlich Ascites Tumor(EAT and Hepatoma AH109A(HAH). Inflammatory tissue induced by turpentine oil was used as an abscess model. Serial scintigraphic images were obtained following IV injections of 100 ..mu..Ci of I-131 or In-111-DTPA-PNA. The tumor affinity of Ga-67 citrate was studied to compare that of radiolabeled PNA. Tissue biodistribution was studied in EAT bearing mice. All of these tumor models except HAH were clearly visible by radiolabeled PNA without subtraction techniques. In the models of LLC and EAT, PNA showed the better accumulation into the tumor tissue than Ga-67 citrate. In YS and MM, PNA represented almost the same accumulation as Ga-67 citrate. The localization of PNA into abscess tissue wasn't found although Ga-67 citrate markedly accumulated into abscess tissue as well as tumor tissue. The clearance of PNA from tumor was slower than those from any other organs. Tumor to muscle ratio was 5.1 at 48hrs. and tumor to blood ratio increased with time to 2.3 at 96hrs. These results suggested that radiolabeled PNA may have a potential in the detection of tumor.

  8. Molecular modeling of oscillating GHz electric field influence on the kinesin affinity to microtubule

    NASA Astrophysics Data System (ADS)

    R. Saeidi, H.; S. Setayandeh, S.; Lohrasebi, A.

    2015-08-01

    Kinesin is a microtubule-associated motor protein which can respond to the external electric field due to its polarity. Using a molecular dynamics simulation method, the effect of such a field on the affinity of kinesin to the αβ-tubulin is investigated in this study. To consider kinesin affinity, the system is exposed to an electric field of 0.03 V/nm with frequency values of 1, 2, …, 9, and 10 GHz. It is found that the applied electric field can change kinesin affinity to the microtubule. These changes could perturb the normal operation of kinesin, such as the processive motility of kinesin on the microtubule.

  9. Constraints on the affinity term for modeling long-term glass dissolution rates

    SciTech Connect

    Bourcier, W.L.; Carroll, S.A.; Phillips, B.L.

    1993-11-01

    Predictions of long-term glass dissolution rates are highly dependent on the form of the affinity term in the rate expression. Analysis of the quantitative effect of saturation state on glass dissolution rate for CSG glass (a simple analog of SRL-165 glass), shows that a simple (1-Q/K) affinity term does not match experimental results. Our data at 100{degree}C show that the data is better fit by an affinity term having the form (1 {minus} (Q/K){sup 1}/{sigma}) where {sigma} = 10.

  10. Energetics of ligand-receptor binding affinity on endothelial cells: An in vitro model.

    PubMed

    Fotticchia, Iolanda; Guarnieri, Daniela; Fotticchia, Teresa; Falanga, Andrea Patrizia; Vecchione, Raffaele; Giancola, Concetta; Netti, Paolo Antonio

    2016-08-01

    Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid and reliable methodology based on Isothermal Titration Calorimetry (ITC) coupled with confluent cell layers cultured around biocompatible templating microparticles to quantify the number of overexpressing receptors on cell membrane and study the energetics of receptor-ligand binding in near-physiological conditions. In the in vitro model here proposed we used the bEnd3 cell line as brain endothelial cells to mimic the blood brain barrier (BBB) cultured on dextran microbeads ranging from 67μm to 80μm in size (Cytodex) and the primary human umbilical vein cells (HUVEC) for comparison. The revealed affinity between transferrin (Tf) and transferrin receptor (TfR) in both systems is very high, Kd values are in the order of nM. Conversely, the value of TfRs/cell reveals a 100-fold increase in the number of TfRs per bEnd3 cells compared to HUVEC cells. The presented methodology can represent a novel and helpful strategy to identify targets, to address drug design and selectively deliver therapeutics that can cross biological barriers such as the blood brain barrier. PMID:27100851

  11. Binding Affinity Effects on Physical Characteristics of a Model Phase-Separated Protein Droplet

    NASA Astrophysics Data System (ADS)

    Chuang, Sara; Banani, Salman; Rosen, Michael; Brangwynne, Clifford

    2015-03-01

    Non-membrane bound organelles are associated with a range of biological functions. Several of these structures exhibit liquid-like properties, and may represent droplets of phase-separated RNA and/or proteins. These structures are often enriched in multi-valent molecules, however little is known about the interactions driving the assembly, properties, and function. Here, we address this question using a model multi-valent protein system consisting of repeats of Small Ubiquitin-like Modifier (SUMO) protein and a SUMO-interacting motif (SIM). These proteins undergo phase separation into liquid-like droplets. We combine microrheology and quantitative microscopy to determine affect of binding affinity on the viscosity, density and surface tension of these droplets. We also use fluorescence recovery after photobleaching (FRAP), fluorescence correlation spectroscopy (FCS) and partitioning experiments to probe the structure and dynamics within these droplets. Our results shed light on how inter-molecular interactions manifests in droplet properties, and lay the groundwork for a comprehensive biophysical picture of intracellular RNA/protein organelles.

  12. Monolayers of a Model Anesthetic-Binding Membrane Protein: Formation, Characterization, and Halothane-Binding Affinity

    PubMed Central

    Churbanova, Inna Y.; Tronin, Andrey; Strzalka, Joseph; Gog, Thomas; Kuzmenko, Ivan; Johansson, Jonas S.; Blasie, J. Kent

    2006-01-01

    hbAP0 is a model membrane protein designed to possess an anesthetic-binding cavity in its hydrophilic domain and a cation channel in its hydrophobic domain. Grazing incidence x-ray diffraction shows that hbAP0 forms four-helix bundles that are vectorially oriented within Langmuir monolayers at the air-water interface. Single monolayers of hbAP0 on alkylated solid substrates would provide an optimal system for detailed structural and dynamical studies of anesthetic-peptide interaction via x-ray and neutron scattering and polarized spectroscopic techniques. Langmuir-Blodgett and Langmuir-Schaeffer deposition and self-assembly techniques were used to form single monolayer films of the vectorially oriented peptide hbAP0 via both chemisorption and physisorption onto suitably alkylated solid substrates. The films were characterized by ultraviolet absorption, ellipsometry, circular dichroism, and polarized Fourier transform infrared spectroscopy. The α-helical secondary structure of the peptide was retained in the films. Under certain conditions, the average orientation of the helical axis was inclined relative to the plane of the substrate, approaching perpendicular in some cases. The halothane-binding affinity of the vectorially oriented hbAP0 peptide in the single monolayers, with the volatile anesthetic introduced into the moist vapor environment of the monolayer, was found to be similar to that for the detergent-solubilized peptide. PMID:16473900

  13. Synthetic cannabinoids: In silico prediction of the cannabinoid receptor 1 affinity by a quantitative structure-activity relationship model.

    PubMed

    Paulke, Alexander; Proschak, Ewgenij; Sommer, Kai; Achenbach, Janosch; Wunder, Cora; Toennes, Stefan W

    2016-03-14

    The number of new synthetic psychoactive compounds increase steadily. Among the group of these psychoactive compounds, the synthetic cannabinoids (SCBs) are most popular and serve as a substitute of herbal cannabis. More than 600 of these substances already exist. For some SCBs the in vitro cannabinoid receptor 1 (CB1) affinity is known, but for the majority it is unknown. A quantitative structure-activity relationship (QSAR) model was developed, which allows the determination of the SCBs affinity to CB1 (expressed as binding constant (Ki)) without reference substances. The chemically advance template search descriptor was used for vector representation of the compound structures. The similarity between two molecules was calculated using the Feature-Pair Distribution Similarity. The Ki values were calculated using the Inverse Distance Weighting method. The prediction model was validated using a cross validation procedure. The predicted Ki values of some new SCBs were in a range between 20 (considerably higher affinity to CB1 than THC) to 468 (considerably lower affinity to CB1 than THC). The present QSAR model can serve as a simple, fast and cheap tool to get a first hint of the biological activity of new synthetic cannabinoids or of other new psychoactive compounds. PMID:26795018

  14. Predicting ligand binding affinity with alchemical free energy methods in a polar model binding site

    PubMed Central

    Boyce, Sarah E.; Mobley, David L.; Rocklin, Gabriel; Graves, Alan P.

    2009-01-01

    We present a combined experimental and modeling study of organic ligand molecules binding to a slightly polar engineered cavity site in T4 lysozyme (L99A/M102Q). For modeling, we computed alchemical absolute binding free energies. These were blind tests performed prospectively on 13 diverse, previously untested candidate ligand molecules. We predicted that eight compounds would bind to the cavity and five would not; 11 of 13 predictions were correct at this level. The RMS error to the measurable absolute binding energies was 1.8 kcal/mol. In addition, we computed relative binding free energies for six phenol derivatives starting from two known ligands: phenol and catechol. The average RMS error in the relative free energy prediction was 2.5 (phenol) and 1.1 (catechol) kcal/mol. To understand these results at atomic resolution, we obtained x-ray co-complex structures for nine of the diverse ligands and for all six phenol analogs. The average RMSD of the predicted pose to the experiment was 2.0Å (diverse set), 1.8Å (phenol derived predictions) and 1.2Å (catechol derived predictions). We found that to predict accurate affinities and rank-orderings required near-native starting orientations of the ligand in the binding site. Unanticipated binding modes, multiple ligand binding, and protein conformational change all proved challenging for the free energy methods. We believe these results can help guide future improvements in physics-based absolute binding free energy methods. PMID:19782087

  15. Diffusion modelling of metamorphic layered coronas with stability criterion and consideration of affinity

    NASA Astrophysics Data System (ADS)

    Ashworth, J. R.; Sheplev, V. S.

    1997-09-01

    Layered coronas between two reactant minerals can, in many cases, be attributed to diffusion-controlled growth with local equilibrium. This paper clarifies and unifies the previous approaches of various authors to the simplest form of modelling, which uses no assumed values for thermochemical quantities. A realistic overall reaction must be estimated from measured overall proportions of minerals and their major element compositions. Modelling is not restricted to a particular number of components S, relative to the number of phases Φ. IfΦ > S + 1, the overall reaction is a combination of simultaneous reactions. The stepwise method, solving for the local reaction at each boundary in turn, is extended to allow for recurrence of a mineral (its presence in two parts of the layer structure separated by a gap). The equations are also given in matrix form. A thermodynamic stability criterion is derived, determining which layer sequence is truly stable if several are computable from the same inputs. A layer structure satisfying the stability criterion has greater growth rate (and greater rate of entropy production) than the other computable layer sequences. This criterion of greatest entropy production is distinct from Prigogine's theorem of minimum entropy production, which distinguishes the stationary or quasi-stationary state from other states of the same layer sequence. The criterion leads to modification of previous results for coronas comprising hornblende, spinel, and orthopyroxene between olivine (Ol) and plagioclase (Pl). The outcome supports the previous inference that Si, and particularly Al, commonly behave as immobile relative to other cation-forming major elements. The affinity (-ΔG) of a corona-forming reaction is estimated, using previous estimates of diffusion coefficient and the duration t of reaction, together with a new model quantity (-ΔG) *. For an example of the Ol + Pl reaction, a rough calculation gives (-ΔG) > 1.7RT (per mole of P1 consumed

  16. Fuzzy controller design for passive continuous-time affine T-S fuzzy models with relaxed stability conditions.

    PubMed

    Chang, Wen-Jer; Ku, Cheung-Chieh; Huang, Pei-Hwa; Chang, Wei

    2009-07-01

    In order to design a fuzzy controller for complex nonlinear systems, the work of this paper deals with developing the relaxed stability conditions for continuous-time affine Takagi-Sugeno (T-S) fuzzy models. By applying the passivity theory and Lyapunov theory, the relaxed stability conditions are derived to guarantee the stability and passivity property of closed-loop systems. Based on these relaxed stability conditions, the synthesis of fuzzy controller design problem for passive continuous-time affine T-S fuzzy models can be easily solved via the Optimal Convex Programming Algorithm (OCPA) and Linear Matrix Inequality (LMI) technique. At last, a simulation example for the fuzzy control of a nonlinear synchronous generator system is presented to manifest the applications and effectiveness of proposed fuzzy controller design approach. PMID:19389667

  17. Affinity of Aporphines for the Human 5-HT2A Receptor: Insights from Homology Modeling and Molecular Docking Studies

    PubMed Central

    Pecic, Stevan; Makkar, Pooja; Chaudhary, Sandeep; Reddy, Boojala V.; Navarro, Hernan A.; Harding, Wayne W.

    2010-01-01

    Analogs of nantenine were docked into a modeled structure of the human 5-HT2A receptor using ICM Pro, GLIDE and GOLD docking methods. The resultant docking scores were used to correlate with observed in vitro apparent affinity (Ke) data. The GOLD docking algorithm when used with a homology model of 5-HT2A, based on a bovine rhodopsin template and built by the program MODELLER, gives results which are most in agreement with the in vitro results. Further analysis of the docking poses among members of a C1 alkyl series of nantenine analogs, indicate that they bind to the receptor in a similar orientation, but differently than nantenine. Besides an important interaction between the protonated nitrogen of the C1 alkyl analogs and residue Asp155, we identified Ser242, Phe234 and Gly238 as key residues responsible for the affinity of these compounds for the 5-HT2A receptor. Specifically, the ability of some of these analogs to establish a H-bond with Ser242 and hydrophobic interactions with Phe234 and Gly238 appears to explain their enhanced affinity as compared to nantenine. PMID:20621490

  18. Toda Molecule Equation and Quotient-Difference Method

    NASA Astrophysics Data System (ADS)

    Sogo, Kiyoshi

    1993-04-01

    The numerical algorithm for computing eigenvalues of a given matrix using the Toda molecule equation, suggested recently by Hirota, Tsujimoto and Imai, is shown to be equivalent to the quotient-difference method. This relation, convergence of the algorithm and extension to a much wider range of matrices are described.

  19. Dispersionless 2D Toda hierarchy, Hurwitz numbers and Riemann theorem

    NASA Astrophysics Data System (ADS)

    Natanzon, Sergey M.

    2016-01-01

    We describe all formal symmetric solutions of dispersionless 2D Toda hierarchy. This classification we use for solving of two classical problems: 1) The calculation of conformal mapping of an arbitrary simply connected domain to the standard disk; 2) Calculation of 2- Hurwitz numbers of genus 0.

  20. Mixed-model QSAR at the glucocorticoid receptor: predicting the binding mode and affinity of psychotropic drugs.

    PubMed

    Spreafico, Morena; Ernst, Beat; Lill, Markus A; Smiesko, Martin; Vedani, Angelo

    2009-01-01

    The glucocorticoid receptor (GR) is a member of the nuclear receptor superfamily that affects immune response, development, and metabolism in target tissues. Glucocorticoids are widely used to treat diverse pathophysiological conditions, but their clinical applicability is limited by side effects. A prediction of the binding affinity toward the GR would be beneficial for identifying glucocorticoid-mediated adverse effects triggered by drugs or chemicals. By identifying the binding mode to the GR using flexible docking (software Yeti) and quantifying the binding affinity through multidimensional QSAR (software Quasar), we validated a model family based on 110 compounds, representing four different chemical classes. The correlation with the experimental data (cross-validated r(2)=0.702; predictive r(2)=0.719) suggests that our approach is suited for predicting the binding affinity of related compounds toward the GR. After challenging the model by a series of scramble tests, a consensus approach (software Raptor), and a prediction set, it was incorporated into our VirtualToxLab and used to simulate and quantify the interaction of 24 psychotropic drugs with the GR. PMID:19009570

  1. Towards Analytic Solutions of Step-Wise Safe Switching for Known Affine-Linear Models

    SciTech Connect

    Koumboulis, Fotis N.; Tzamtzi, Maria P.

    2008-09-17

    In the present work we establish conditions which guarantee safe transitions for the closed-loop system produced by the application of the Step-Wise Safe Switching control approach to an affine linear system when the nonlinear description of the plant is known. These conditions are based on the local Input to State Stability (ISS) properties of the nonlinear system around the plant's nominal operating points.

  2. Affinity of HIV-1 antibody 2G12 with monosaccharides: a theoretical study based on explicit and implicit water models.

    PubMed

    Koyama, Yuka; Ueno-Noto, Kaori; Takano, Keiko

    2014-04-01

    In order to develop potential ligands to HIV-1 antibody 2G12 toward HIV-1 vaccine, binding mechanisms of the antibody 2G12 with the glycan ligand of D-mannose and D-fructose were theoretically examined. D-Fructose, whose molecular structure is slightly different from D-mannose, has experimentally shown to have stronger binding affinity to the antibody than that of D-mannose. To clarify the nature of D-fructose's higher binding affinity over D-mannose, we studied interaction between the monosaccharides and the antibody using ab initio fragment molecular orbital (FMO) method considering solvation effect as implicit model (FMO-PCM) as well as explicit water model. The calculated binding free energies of the glycans were qualitatively well consistent with the experimentally reported order of their affinities with the antibody 2G12. In addition, the FMO-PCM calculation elucidated the advantages of D-fructose over D-mannose in the solvation energy as well as the entropic contribution term obtained by MD simulations. The effects of explicit water molecules observed in the X-ray crystal structure were also scrutinized by means of FMO methods. Significant pair interaction energies among D-fructose, amino acids, and water molecules were uncovered, which indicated contributions from the water molecules to the strong binding ability of D-fructose to the antibody 2G12. These FMO calculation results of explicit water model as well as implicit water model indicated that the strong binding of D-fructose over D-mannose was due to the solvation effects on the D-fructose interaction energy. PMID:24583603

  3. q-bosons, Toda lattice, Pieri rules and Baxter q-operator

    NASA Astrophysics Data System (ADS)

    Duval, Antoine; Pasquier, Vincent

    2016-04-01

    We use the Pieri rules to recover the q-boson model and show it is equivalent to a discretized version of the relativistic Toda chain. We identify its semi infinite transfer matrix and the corresponding Baxter Q-matrix with half vertex operators related by an ω-duality transformation. We observe that the scalar product of two higher spin XXZ wave functions can be expressed with a Gaudin determinant. In honour of Rodney Baxter’s 75th birthday.

  4. Towards the chemometric dissection of peptide--HLA-A*0201 binding affinity: comparison of local and global QSAR models.

    PubMed

    Doytchinova, Irini A; Walshe, Valerie; Borrow, Persephone; Flower, Darren R

    2005-03-01

    The affinities of 177 nonameric peptides binding to the HLA-A*0201 molecule were measured using a FACS-based MHC stabilisation assay and analysed using chemometrics. Their structures were described by global and local descriptors, QSAR models were derived by genetic algorithm, stepwise regression and PLS. The global molecular descriptors included molecular connectivity chi indices, kappa shape indices, E-state indices, molecular properties like molecular weight and log P, and three-dimensional descriptors like polarizability, surface area and volume. The local descriptors were of two types. The first used a binary string to indicate the presence of each amino acid type at each position of the peptide. The second was also position-dependent but used five z-scales to describe the main physicochemical properties of the amino acids forming the peptides. The models were developed using a representative training set of 131 peptides and validated using an independent test set of 46 peptides. It was found that the global descriptors could not explain the variance in the training set nor predict the affinities of the test set accurately. Both types of local descriptors gave QSAR models with better explained variance and predictive ability. The results suggest that, in their interactions with the MHC molecule, the peptide acts as a complicated ensemble of multiple amino acids mutually potentiating each other. PMID:16059672

  5. Prediction of binding affinity and efficacy of thyroid hormone receptor ligands using QSAR and structure-based modeling methods

    SciTech Connect

    Politi, Regina; Rusyn, Ivan; Tropsha, Alexander

    2014-10-01

    The thyroid hormone receptor (THR) is an important member of the nuclear receptor family that can be activated by endocrine disrupting chemicals (EDC). Quantitative Structure–Activity Relationship (QSAR) models have been developed to facilitate the prioritization of THR-mediated EDC for the experimental validation. The largest database of binding affinities available at the time of the study for ligand binding domain (LBD) of THRβ was assembled to generate both continuous and classification QSAR models with an external accuracy of R{sup 2} = 0.55 and CCR = 0.76, respectively. In addition, for the first time a QSAR model was developed to predict binding affinities of antagonists inhibiting the interaction of coactivators with the AF-2 domain of THRβ (R{sup 2} = 0.70). Furthermore, molecular docking studies were performed for a set of THRβ ligands (57 agonists and 15 antagonists of LBD, 210 antagonists of the AF-2 domain, supplemented by putative decoys/non-binders) using several THRβ structures retrieved from the Protein Data Bank. We found that two agonist-bound THRβ conformations could effectively discriminate their corresponding ligands from presumed non-binders. Moreover, one of the agonist conformations could discriminate agonists from antagonists. Finally, we have conducted virtual screening of a chemical library compiled by the EPA as part of the Tox21 program to identify potential THRβ-mediated EDCs using both QSAR models and docking. We concluded that the library is unlikely to have any EDC that would bind to the THRβ. Models developed in this study can be employed either to identify environmental chemicals interacting with the THR or, conversely, to eliminate the THR-mediated mechanism of action for chemicals of concern. - Highlights: • This is the largest curated dataset for ligand binding domain (LBD) of the THRβ. • We report the first QSAR model for antagonists of AF-2 domain of THRβ. • A combination of QSAR and docking enables

  6. Nonlinear dust-lattice waves: a modified Toda lattice

    SciTech Connect

    Cramer, N. F.

    2008-09-07

    Charged dust grains in a plasma interact with a Coulomb potential, but also with an exponential component to the potential, due to Debye shielding in the background plasma. Here we investigate large-amplitude oscillations and waves in dust-lattices, employing techniques used in Toda lattice analysis. The lattice consists of a linear chain of particles, or a periodic ring as occurs in experimentally observed dust particle clusters. The particle motion has a triangular waveform, and chaotic motion for large amplitude motion of a grain.

  7. On the squared eigenfunction symmetry of the Toda lattice hierarchy

    NASA Astrophysics Data System (ADS)

    Cheng, Jipeng; He, Jingsong

    2013-02-01

    The squared eigenfunction symmetry for the Toda lattice hierarchy is explicitly constructed in the form of the Kronecker product of the vector eigenfunction and the vector adjoint eigenfunction, which can be viewed as the generating function for the additional symmetries when the eigenfunction and the adjoint eigenfunction are the wave function and the adjoint wave function, respectively. Then after the Fay-like identities and some important relations about the wave functions are investigated, the action of the squared eigenfunction related to the additional symmetry on the tau function is derived, which is equivalent to the Adler-Shiota-van Moerbeke formulas.

  8. Exact quantization conditions for the relativistic Toda lattice

    NASA Astrophysics Data System (ADS)

    Hatsuda, Yasuyuki; Mariño, Marcos

    2016-05-01

    Inspired by recent connections between spectral theory and topological string theory, we propose exact quantization conditions for the relativistic Toda lattice of N particles. These conditions involve the Nekrasov-Shatashvili free energy, which resums the perturbative WKB expansion, but they require in addition a non-perturbative contribution, which is related to the perturbative result by an S-duality transformation of the Planck constant. We test the quantization conditions against explicit calculations of the spectrum for N = 3. Our proposal can be generalized to arbitrary toric Calabi-Yau manifolds and might solve the corresponding quantum integrable system of Goncharov and Kenyon.

  9. Prediction of binding affinity and efficacy of thyroid hormone receptor ligands using QSAR and structure based modeling methods

    PubMed Central

    Politi, Regina; Rusyn, Ivan; Tropsha, Alexander

    2016-01-01

    The thyroid hormone receptor (THR) is an important member of the nuclear receptor family that can be activated by endocrine disrupting chemicals (EDC). Quantitative Structure-Activity Relationship (QSAR) models have been developed to facilitate the prioritization of THR-mediated EDC for the experimental validation. The largest database of binding affinities available at the time of the study for ligand binding domain (LBD) of THRβ was assembled to generate both continuous and classification QSAR models with an external accuracy of R2=0.55 and CCR=0.76, respectively. In addition, for the first time a QSAR model was developed to predict binding affinities of antagonists inhibiting the interaction of coactivators with the AF-2 domain of THRβ (R2=0.70). Furthermore, molecular docking studies were performed for a set of THRβ ligands (57 agonists and 15 antagonists of LBD, 210 antagonists of the AF-2 domain, supplemented by putative decoys/non-binders) using several THRβ structures retrieved from the Protein Data Bank. We found that two agonist-bound THRβ conformations could effectively discriminate their corresponding ligands from presumed non-binders. Moreover, one of the agonist conformations could discriminate agonists from antagonists. Finally, we have conducted virtual screening of a chemical library compiled by the EPA as part of the Tox21 program to identify potential THRβ-mediated EDCs using both QSAR models and docking. We concluded that the library is unlikely to have any EDC that would bind to the THRβ. Models developed in this study can be employed either to identify environmental chemicals interacting with the THR or, conversely, to eliminate the THR-mediated mechanism of action for chemicals of concern. PMID:25058446

  10. Multifold Darboux Transformations of the Extended Bigraded Toda Hierarchy

    NASA Astrophysics Data System (ADS)

    Li, Chuanzhong; Song, Tao

    2016-04-01

    With the extended logarithmic flow equations and some extended Vertex operators in generalized Hirota bilinear equations, extended bigraded Toda hierarchy (EBTH) was proved to govern the Gromov-Witten theory of orbiford cNM in literature. The generating function of these Gromov-Witten invariants is one special solution of the EBTH. In this article, the multifold Darboux transformations and their determinant representations of the EBTH are given with two different gauge transformation operators. The two Darboux transformations in different directions are used to generate new solutions from known solutions which include soliton solutions of (N, N)-EBTH, i.e. the EBTH when N=M. From the generation of new solutions, we can find the big difference between the EBTH and the extended Toda hierarchy (ETH). Also, we plotted the soliton graphs of the (N, N)-EBTH from which some approximation analysis is given. From the analysis on velocities of soliton solutions, the difference between the extended flows and other flows are shown. The two different Darboux transformations constructed by us might be useful in Gromov-Witten theory of orbiford cNM.

  11. An affine continuum mechanical model for cross-linked F-actin networks with compliant linker proteins.

    PubMed

    Holzapfel, Gerhard A; Unterberger, Michael J; Ogden, Ray W

    2014-10-01

    Cross-linked actin networks are important building blocks of the cytoskeleton. In order to gain deeper insight into the interpretation of experimental data on actin networks, adequate models are required. In this paper we introduce an affine constitutive network model for cross-linked F-actin networks based on nonlinear continuum mechanics, and specialize it in order to reproduce the experimental behavior of in vitro reconstituted model networks. The model is based on the elastic properties of single filaments embedded in an isotropic matrix such that the overall properties of the composite are described by a free-energy function. In particular, we are able to obtain the experimentally determined shear and normal stress responses of cross-linked actin networks typically observed in rheometer tests. In the present study an extensive analysis is performed by applying the proposed model network to a simple shear deformation. The single filament model is then extended by incorporating the compliance of cross-linker proteins and further extended by including viscoelasticity. All that is needed for the finite element implementation is the constitutive model for the filaments, the linkers and the matrix, and the associated elasticity tensor in either the Lagrangian or Eulerian formulation. The model facilitates parameter studies of experimental setups such as micropipette aspiration experiments and we present such studies to illustrate the efficacy of this modeling approach. PMID:25043658

  12. Robust fuzzy output feedback controller for affine nonlinear systems via T-S fuzzy bilinear model: CSTR benchmark.

    PubMed

    Hamdy, M; Hamdan, I

    2015-07-01

    In this paper, a robust H∞ fuzzy output feedback controller is designed for a class of affine nonlinear systems with disturbance via Takagi-Sugeno (T-S) fuzzy bilinear model. The parallel distributed compensation (PDC) technique is utilized to design a fuzzy controller. The stability conditions of the overall closed loop T-S fuzzy bilinear model are formulated in terms of Lyapunov function via linear matrix inequality (LMI). The control law is robustified by H∞ sense to attenuate external disturbance. Moreover, the desired controller gains can be obtained by solving a set of LMI. A continuous stirred tank reactor (CSTR), which is a benchmark problem in nonlinear process control, is discussed in detail to verify the effectiveness of the proposed approach with a comparative study. PMID:25765955

  13. Failure of the three compartment model to describe the pharmacokinetics in brain of a high affinity substituted benzamide.

    PubMed

    Votaw, J R; Kessler, R M; de Paulis, T

    1993-11-01

    The applicability of using the standard 3-compartment model to describe the neuropharmacokinetics of a high affinity substituted benzamide was investigated. We performed the following experiments using the [18F]-5-(3-fluoropropyl) analog of epidepride ([18F]5-FPrEpid), a potent dopamine D2 receptor antagonist: constant left ventricular infusion, first-pass clearance, varying ligand specific activity, and displacing bound ligand with varying amounts of unlabelled ligand. Taken together, the information from these experiments rigorously tests the standard 3-compartment model. The obtained data and predictions from the model of the kinetic behavior of the ligand are inconsistent. The measured and model predicted dissociation rate (measured koff = 0.065 min-1, model prediction koff = 0.007 min-1) and the equilibrium dissociation constant (measured KD = 0.14 nM, model prediction KD = 2.2 nM) differ by an order of magnitude. Furthermore, the model cannot be used to accurately estimate the receptor density. We postulate that the synapse geometry and physical relationship between receptors are necessary components of a model that describes the pharmacokinetics of [18F]5-FPrEpid. PMID:8278896

  14. Low oxygen affinity in reptilian hemoglobin D: prediction of residue interactions in Geochelone carbonaria HbD by homology modeling.

    PubMed

    Lutfullah, Ghosia; Khalil, Hilal Shahid; Amin, Farhat; Azhar, Noreen

    2008-04-01

    The homology model of hemoglobin D from Geochelone carbonaria, the red-footed tortoise was predicted using the 3D structure coordinates of Geochelone gigantea hemoglobin D as the template. The model was built using the program, MODELLER (8v1) and evaluated with PROCHECK and PROSA. The present study features an in-depth analysis of the 3D model and its conformational changes brought about with variations in its environment. These structural changes are correlated with its ability to adapt to different environmental constraints enabling the organism to better suit to its natural habitat. The model shows additional contacts between amino acid pairs of alpha-119 and beta-55, alpha-35 and beta-124, alpha-103 and beta-112, alpha-115 and beta-116, alpha-120 and beta-52, alpha-120 and beta-55, alpha-36 and beta-127 which are not found in human hemoglobin. It is predicted that these contacts may result in T-state stabilization thus lowering oxygen affinity. Furthermore, decrease in the interaction of phosphate heteroatoms with the amino acid residues of G. carbonaria Hb was also predicted in this study. PMID:18085430

  15. Modeling Taylor dispersion injections: determination of kinetic/affinity interaction constants and diffusion coefficients in label-free biosensing.

    PubMed

    Quinn, John G

    2012-02-15

    A new method based on Taylor dispersion has been developed that enables an analyte gradient to be titrated over a ligand-coated surface for kinetic/affinity analysis of interactions from a minimal number of injections. Taylor dispersion injections generate concentration ranges in excess of four orders of magnitude and enable the analyte diffusion coefficient to be reliably estimated as a fitted parameter when fitting binding interaction models. A numerical model based on finite element analysis, Monte Carlo simulations, and statistical profiling were used to compare the Taylor dispersion method with standard fixed concentration injections in terms of parameter correlation, linearity of parameter error space, and global versus local model fitting. A dramatic decrease in parameter correlations was observed for TDi curves relative to curves from standard fixed concentration injections when surface saturation was achieved. In FCI the binding progress is recorded with respect to injection time, whereas in TDi the second time dependency encoded in the analyte gradient increases resolving power. This greatly lowers the dependence of all parameters on each other and on experimental interferences. When model parameters were fitted locally, the performance of TDis remained comparable to global model fitting, whereas fixed concentration binding response curves yielded unreliable parameter estimates. PMID:22197421

  16. Mathematical Model of the Firefly Luciferase Complementation Assay Reveals a Non-Linear Relationship between the Detected Luminescence and the Affinity of the Protein Pair Being Analyzed.

    PubMed

    Dale, Renee; Ohmuro-Matsuyama, Yuki; Ueda, Hiroshi; Kato, Naohiro

    2016-01-01

    The firefly luciferase complementation assay is widely used as a bioluminescent reporter technology to detect protein-protein interactions in vitro, in cellulo, and in vivo. Upon the interaction of a protein pair, complemented firefly luciferase emits light through the adenylation and oxidation of its substrate, luciferin. Although it has been suggested that kinetics of light production in the firefly luciferase complementation assay is different from that in full length luciferase, the mechanism behind this is still not understood. To quantitatively understand the different kinetics and how changes in affinity of a protein pair affect the light emission in the assay, a mathematical model of the in vitro firefly luciferase complementation assay was constructed. Analysis of the model finds that the change in kinetics is caused by rapid dissociation of the protein pair, low adenylation rate of luciferin, and increased affinity of adenylated luciferin to the enzyme. The model suggests that the affinity of the protein pair has an exponential relationship with the light detected in the assay. This relationship causes the change of affinity in a protein pair to be underestimated. This study underlines the importance of understanding the molecular mechanism of the firefly luciferase complementation assay in order to analyze protein pair affinities quantitatively. PMID:26886551

  17. Mathematical Model of the Firefly Luciferase Complementation Assay Reveals a Non-Linear Relationship between the Detected Luminescence and the Affinity of the Protein Pair Being Analyzed

    PubMed Central

    Dale, Renee; Ohmuro-Matsuyama, Yuki; Ueda, Hiroshi; Kato, Naohiro

    2016-01-01

    The firefly luciferase complementation assay is widely used as a bioluminescent reporter technology to detect protein-protein interactions in vitro, in cellulo, and in vivo. Upon the interaction of a protein pair, complemented firefly luciferase emits light through the adenylation and oxidation of its substrate, luciferin. Although it has been suggested that kinetics of light production in the firefly luciferase complementation assay is different from that in full length luciferase, the mechanism behind this is still not understood. To quantitatively understand the different kinetics and how changes in affinity of a protein pair affect the light emission in the assay, a mathematical model of the in vitro firefly luciferase complementation assay was constructed. Analysis of the model finds that the change in kinetics is caused by rapid dissociation of the protein pair, low adenylation rate of luciferin, and increased affinity of adenylated luciferin to the enzyme. The model suggests that the affinity of the protein pair has an exponential relationship with the light detected in the assay. This relationship causes the change of affinity in a protein pair to be underestimated. This study underlines the importance of understanding the molecular mechanism of the firefly luciferase complementation assay in order to analyze protein pair affinities quantitatively. PMID:26886551

  18. Report: Affinity Chromatography.

    ERIC Educational Resources Information Center

    Walters, Rodney R.

    1985-01-01

    Supports, affinity ligands, immobilization, elution methods, and a number of applications are among the topics considered in this discussion of affinity chromatography. An outline of the basic principles of affinity chromatography is included. (JN)

  19. Mixed-model QSAR at the human mineralocorticoid receptor: predicting binding mode and affinity of anabolic steroids.

    PubMed

    Peristera, Ourania; Spreafico, Morena; Smiesko, Martin; Ernst, Beat; Vedani, Angelo

    2009-09-28

    We present a computational study on the human mineralocorticoid receptor (hMR) that is based on multi-dimensional quantitative structure-activity relationships (mQSAR). Therein, we identified the binding mode of 48 steroid and non-steroid homologues by flexible docking to the crystal structure (software Yeti) and quantified it using 6D-QSAR (software Quasar). The receptor surrogate, evolved using a genetic algorithm, converged at a cross-validated r2 of 0.810, and yielded a predictive r2 of 0.661. The model was challenged by a series of scramble tests and by consensus scoring (software Raptor: r2=0.844, predictive r(2)=0.620). The model was then employed to predict the binding affinity of 26 anabolic steroids, demonstrating to which extent they might disrupt the endocrine system via binding to the hMR. The model for the hMR was added to the VirtualToxLab, a technology developed by the Biographics Laboratory 3R, allows the identification of the endocrine-disrupting potential of drugs, chemicals and natural products in silico. PMID:19523507

  20. From the affinity constant to the half-saturation index: understanding conventional modeling concepts in novel wastewater treatment processes.

    PubMed

    Arnaldos, Marina; Amerlinck, Youri; Rehman, Usman; Maere, Thomas; Van Hoey, Stijn; Naessens, Wouter; Nopens, Ingmar

    2015-03-01

    The "affinity constant" (KS) concept is applied in wastewater treatment models to incorporate the effect of substrate limitation on process performance. As an increasing number of wastewater treatment processes rely on low substrate concentrations, a proper understanding of these so-called constants is critical in order to soundly model and evaluate emerging treatment systems. In this paper, an in-depth analysis of the KS concept has been carried out, focusing on the different physical and biological phenomena that affect its observed value. By structuring the factors influencing half-saturation indices (newly proposed nomenclature) into advectional, diffusional and biological, light has been shed onto some of the apparent inconsistencies present in the literature. Particularly, the importance of non-ideal mixing as a source of variability between observed KS values in different systems has been illustrated. Additionally, discussion on the differences existent between substrates that affect half-saturation indices has been carried out; it has been shown that the observed KS for some substrates will reflect transport or biological limitations more than others. Finally, potential modeling strategies that could alleviate the shortcomings of the KS concept have been provided. These could be of special importance when considering the evaluation and design of emerging wastewater treatment processes. PMID:25576693

  1. Fast Modeling of Binding Affinities by Means of Superposing Significant Interaction Rules (SSIR) Method.

    PubMed

    Besalú, Emili

    2016-01-01

    The Superposing Significant Interaction Rules (SSIR) method is described. It is a general combinatorial and symbolic procedure able to rank compounds belonging to combinatorial analogue series. The procedure generates structure-activity relationship (SAR) models and also serves as an inverse SAR tool. The method is fast and can deal with large databases. SSIR operates from statistical significances calculated from the available library of compounds and according to the previously attached molecular labels of interest or non-interest. The required symbolic codification allows dealing with almost any combinatorial data set, even in a confidential manner, if desired. The application example categorizes molecules as binding or non-binding, and consensus ranking SAR models are generated from training and two distinct cross-validation methods: leave-one-out and balanced leave-two-out (BL2O), the latter being suited for the treatment of binary properties. PMID:27240346

  2. Fast Modeling of Binding Affinities by Means of Superposing Significant Interaction Rules (SSIR) Method

    PubMed Central

    Besalú, Emili

    2016-01-01

    The Superposing Significant Interaction Rules (SSIR) method is described. It is a general combinatorial and symbolic procedure able to rank compounds belonging to combinatorial analogue series. The procedure generates structure-activity relationship (SAR) models and also serves as an inverse SAR tool. The method is fast and can deal with large databases. SSIR operates from statistical significances calculated from the available library of compounds and according to the previously attached molecular labels of interest or non-interest. The required symbolic codification allows dealing with almost any combinatorial data set, even in a confidential manner, if desired. The application example categorizes molecules as binding or non-binding, and consensus ranking SAR models are generated from training and two distinct cross-validation methods: leave-one-out and balanced leave-two-out (BL2O), the latter being suited for the treatment of binary properties. PMID:27240346

  3. New analogues of epiboxidine incorporating the 4,5-dihydroisoxazole nucleus: synthesis, binding affinity at neuronal nicotinic acetylcholine receptors, and molecular modeling investigations.

    PubMed

    Dallanoce, Clelia; Magrone, Pietro; Bazza, Paola; Grazioso, Giovanni; Rizzi, Luca; Riganti, Loredana; Gotti, Cecilia; Clementi, Francesco; Frydenvang, Karla; De Amici, Marco

    2009-02-01

    A group of novel 4,5-dihydro-3-methylisoxazolyl derivatives, structurally related to epiboxidine (=(1R,4S,6S)-6-(3-methylisoxazol-5-yl)-7-azabicyclo[2.2.1]heptane), was prepared via 1,3-dipolar cycloaddition of acetonitrile oxide to different olefins. Target compounds 1a and 1b, 2a and 2b, 3, 4, and 5 were tested for affinity at neuronal nicotinic heteromeric (alpha4beta2) and homomeric (alpha7) acetylcholine receptors. Notably, diastereoisomers 1a and 1b were characterized by a massive drop of the affinity at the alpha4beta2 subtypes (K(i) values spanning the range 4.3-126 microM), when compared with that of epiboxidine (K(i)=0.6 nM). Therefore, the replacement of the 3-methylisoxazole ring of epiboxidine with the 4,5-dihydro-3-methylisoxazole nucleus is detrimental for the affinity at alpha4beta2 receptors. A comparable lack of affinity/selectivity for the two nAChR subtypes under study was evidenced for the remaining epiboxidine-related dihydroisoxazole derivatives 2a and 2b, and 3-5. Diastereoisomers 1a and 1b, and spirocyclic derivative 3 were docked into molecular models of the receptor subtypes under study, and their binding mode was compared with that of reference ligands endowed with high binding affinity. PMID:19235154

  4. Synthesis, structure-affinity relationships, and molecular modeling studies of novel pyrazolo[3,4-c]quinoline derivatives as adenosine receptor antagonists.

    PubMed

    Lenzi, Ombretta; Colotta, Vittoria; Catarzi, Daniela; Varano, Flavia; Squarcialupi, Lucia; Filacchioni, Guido; Varani, Katia; Vincenzi, Fabrizio; Borea, Pier Andrea; Dal Ben, Diego; Lambertucci, Catia; Cristalli, Gloria

    2011-06-15

    This paper reports the study of new 2-phenyl- and 2-methylpyrazolo[3,4-c]quinolin-4-ones (series A) and 4-amines (series B), designed as adenosine receptor (AR) antagonists. The synthesized compounds bear at the 6-position various groups, with different lipophilicity and steric hindrance, that were thought to increase human A(1) and A(2A) AR affinities and selectivities, with respect to those of the parent 6-unsubstituted compounds. In series A, this modification was not tolerated since it reduced AR affinity, while in series B it shifted the binding towards the hA(1) subtype. To rationalize the observed structure-affinity relationships, molecular docking studies at A(2A)AR-based homology models of the A(1) and A(3) ARs and at the A(2A)AR crystal structure were carried out. PMID:21616671

  5. Vesicular stomatitis virus polymerase's strong affinity to its template suggests exotic transcription models.

    PubMed

    Tang, Xiaolin; Bendjennat, Mourad; Saffarian, Saveez

    2014-12-01

    Vesicular stomatitis virus (VSV) is the prototype for negative sense non segmented (NNS) RNA viruses which include potent human and animal pathogens such as Rabies, Ebola and measles. The polymerases of NNS RNA viruses only initiate transcription at or near the 3' end of their genome template. We measured the dissociation constant of VSV polymerases from their whole genome template to be 20 pM. Given this low dissociation constant, initiation and sustainability of transcription becomes nontrivial. To explore possible mechanisms, we simulated the first hour of transcription using Monte Carlo methods and show that a one-time initial dissociation of all polymerases during entry is not sufficient to sustain transcription. We further show that efficient transcription requires a sliding mechanism for non-transcribing polymerases and can be realized with different polymerase-polymerase interactions and distinct template topologies. In conclusion, we highlight a model in which collisions between transcribing and sliding non-transcribing polymerases result in release of the non-transcribing polymerases allowing for redistribution of polymerases between separate templates during transcription and suggest specific experiments to further test these mechanisms. PMID:25501005

  6. Stability, metabolism and transport of D-Asp(OBzl)-Ala--a model prodrug with affinity for the oligopeptide transporter.

    PubMed

    Steffansen, B; Lepist, E I; Taub, M E; Larsen, B D; Frokjaer, S; Lennernäs, H

    1999-04-01

    The model prodrug D-Asp(OBzl)-Ala has previously been shown to have affinity and to be transported by the oligopeptide transporter PepT1 expressed in Caco-2 cells. The main objective of the present study was to investigate the aqueous stability of D-Asp(OBzl)-Ala and its in vitro metabolism in different gastrointestinal media arising from rats and humans, as well as in human plasma. The second major aim of the study was to evaluate our previous study in Caco-2 cell culture, by determining the effective intestinal permeability (Peff) of D-Asp(OBzl)-Ala in situ using the single-pass rat perfusion model. The aqueous stability studies show water, general buffer, as well as specific acid and base catalysis of D-Asp(OBzl)-Ala. The degradation of the model prodrug was independent of ionic strength. The half-lives in rat jejunal fluid and homogenate were >3 h. In human gastric and intestinal fluids, the half-lives were >3 h and 2.3+/-0. 03 h, respectively. Using the rat single-pass perfusion technique, the effective jejunal permeability (Peff) of D-Asp(OBzl)-Ala was determined to be high (1.29+/-0.5.10-4 cm/s). The 32 times higher Peff value found in the perfusion model compared to Caco-2 cells is most likely due to a higher functional expression of the oligopeptide transporter. Rat jejuna Peff was reduced by approximately 50% in the presence of well known oligopeptide transporter substrates, such as Gly-Sar and cephalexin. It may be that D-Asp(OBzl)-Ala is primarily absorbed intact by the rat jejunal oligopeptide transporter, since the stability in the intestinal homogenate and fluids was rather high (t1/2>2.3 h). PMID:10072480

  7. On Differential form Method to Find Lie Symmetries of two Types of Toda Lattices

    NASA Astrophysics Data System (ADS)

    Ding, Qi; Tian, Shou-Fu

    2014-12-01

    In this paper, we investigate Lie symmetries of the (1 + 1)-dimensional celebrated Toda lattice and the (2 + 1)-dimensional modified semidiscrete Toda lattice by using the extended Harrison and Estabrook's geometric approach. Two closed ideals written in terms of a set of differential forms are constructed for Toda lattices. Moreover, commutation relations of a Kac-Moody-Virasoro type Lie algebra are obtained by direct computation.

  8. Mineralogy of new Antarctic achondrites with affinity to Lodran and a model of their evolution in an asteroid

    NASA Technical Reports Server (NTRS)

    Takeda, Hiroshi; Mori, Hiroshi; Hiroi, Takahiro; Saito, Jun

    1994-01-01

    We studied five new Antartic achondrites, MacAlpine Hills (MAC) 88177, Yamato (Y)74357, Y75274, Y791491 and Elephant Moraine (EET)84302 by mineralogical techniques to gain a better understanding of the mineral assemblages of a group of meteorites with an affinity to Lodran (stony-iron meteorite) and their formation processes. This group is being called lodranites. These meteorites contain major coarse-grained orthopyroxene (Opx) and olivine as in Lodran and variable amounts of FeNi metal and troilite etc. MAC88177 has more augite and less FeNi than Lodran; Y74357 has more olivine and contains minor augite; Y791491 contains in addition plagioclase. EET84302 has an Acapulco-like chondritic mineral assembladge and is enriched in FeNi metal and plagioclase, but one part is enriched in Opx and chromite. The EET84302 and MAC88177 Opx crystals have dusty cores as in Acapulco. EET84302 and Y75274 are more Mg-rich than other members of the lodranite group, and Y74357 is intermediate. Since these meteorites all have coarse-grained textures, similar major mineral assemblages, variable amounts of augite, plagioclase, FeNi metal, chromite and olivine, we suggest that they are related and are linked to a parent body with modified chondritic compositions. The variability of the abundances of these minerals are in line with a proposed model of the surface mineral assemblages of the S asteroids. The mineral assemblages can best be explained by differing degrees of loss or movements of lower temperature partial melts and recrystallization, and reduction. A portion of EET84302 rich in metal and plagioclase may represent a type of component removed from the lodranite group meteorites. Y791058 and Caddo County, which were studied for comparison, are plagioclase-rich silicate inclusions in IAB iron meteorites and may have been derived by similar process but in a different body.

  9. A Novel Humanized GLP-1 Receptor Model Enables Both Affinity Purification and Cre-LoxP Deletion of the Receptor

    PubMed Central

    Jun, Lucy S.; Showalter, Aaron D.; Ali, Nosher; Dai, Feihan; Ma, Wenzhen; Coskun, Tamer; Ficorilli, James V.; Wheeler, Michael B.; Michael, M. Dodson; Sloop, Kyle W.

    2014-01-01

    Class B G protein-coupled receptors (GPCRs) are important regulators of endocrine physiology, and peptide-based therapeutics targeting some of these receptors have proven effective at treating disorders such as hypercalcemia, osteoporosis, and type 2 diabetes mellitus (T2DM). As next generation efforts attempt to develop novel non-peptide, orally available molecules for these GPCRs, new animal models expressing human receptor orthologs may be required because small molecule ligands make fewer receptor contacts, and thus, the impact of amino acid differences across species may be substantially greater. The objective of this report was to generate and characterize a new mouse model of the human glucagon-like peptide-1 receptor (hGLP-1R), a class B GPCR for which established peptide therapeutics exist for the treatment of T2DM. hGLP-1R knock-in mice express the receptor from the murine Glp-1r locus. Glucose tolerance tests and gastric emptying studies show hGLP-1R mice and their wild-type littermates display similar physiological responses for glucose metabolism, insulin secretion, and gastric transit, and treatment with the GLP-1R agonist, exendin-4, elicits similar responses in both groups. Further, ex vivo assays show insulin secretion from humanized islets is glucose-dependent and enhanced by GLP-1R agonists. To enable additional utility, the targeting construct of the knock-in line was engineered to contain both flanking LoxP sites and a C-terminal FLAG epitope. Anti-FLAG affinity purification shows strong expression of hGLP-1R in islets, lung, and stomach. We crossed the hGLP-1R line with Rosa26Cre mice and generated global Glp-1r−/− animals. Immunohistochemistry of pancreas from humanized and knock-out mice identified a human GLP-1R-specific antibody that detects the GLP-1R in human pancreas as well as in the pancreas of hGLP-1r knock-in mice. This new hGLP-1R model will allow tissue-specific deletion of the GLP-1R, purification of potential GLP-1R partner

  10. RNase One Gene Isolation, Expression, and Affinity Purification Models Research Experimental Progression and Culminates with Guided Inquiry-Based Experiments

    ERIC Educational Resources Information Center

    Bailey, Cheryl P.

    2009-01-01

    This new biochemistry laboratory course moves through a progression of experiments that generates a platform for guided inquiry-based experiments. RNase One gene is isolated from prokaryotic genomic DNA, expressed as a tagged protein, affinity purified, and tested for activity and substrate specificity. Student pairs present detailed explanations…

  11. A comparative study of lectin affinity based plant n-glycoproteome profiling using tomato fruit as a model

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lectin affinity chromatography (LAC) can provide a valuable front-end enrichment strategy for the study of N-glycoproteins and has been used to characterize a broad range eukaryotic N-glycoproteomes. Moreover, studies with mammalian systems have suggested that the use of multiple lectins with differ...

  12. A Comparative Study of Lectin Affinity Based Plant N-Glycoproteome Profiling Using Tomato Fruit as a Model*

    PubMed Central

    Ruiz-May, Eliel; Hucko, Simon; Howe, Kevin J.; Zhang, Sheng; Sherwood, Robert W.; Thannhauser, Theodore W.; Rose, Jocelyn K. C.

    2014-01-01

    Lectin affinity chromatography (LAC) can provide a valuable front-end enrichment strategy for the study of N-glycoproteins and has been used to characterize a broad range eukaryotic N-glycoproteomes. Moreover, studies with mammalian systems have suggested that the use of multiple lectins with different affinities can be particularly effective. A multi-lectin approach has also been reported to provide a significant benefit for the analysis of plant N-glycoproteins; however, it has yet to be determined whether certain lectins, or combinations of lectins are optimal for plant N-glycoproteome profiling; or whether specific lectins show preferential association with particular N-glycosylation sites or N-glycan structures. We describe here a comparative study of three mannose-binding lectins, concanavalin A, snowdrop lectin, and lentil lectin, to profile the N-glycoproteome of mature green stage tomato (Solanum lycopersicum) fruit pericarp. Through coupling lectin affinity chromatography with a shotgun proteomics strategy, we identified 448 putative N-glycoproteins, whereas a parallel lectin affinity chromatography plus hydrophilic interaction chromatography analysis revealed 318 putative N-glycosylation sites on 230 N-glycoproteins, of which 100 overlapped with the shotgun analysis, as well as 17 N-glycan structures. The use of multiple lectins substantially increased N-glycoproteome coverage and although there were no discernible differences in the structures of N-glycans, or the charge, isoelectric point (pI) or hydrophobicity of the glycopeptides that differentially bound to each lectin, differences were observed in the amino acid frequency at the −1 and +1 subsites of the N-glycosylation sites. We also demonstrated an alternative and complementary in planta recombinant expression strategy, followed by affinity MS analysis, to identify the putative N-glycan structures of glycoproteins whose abundance is too low to be readily determined by a shotgun approach, and

  13. Template CoMFA Generates Single 3D-QSAR Models that, for Twelve of Twelve Biological Targets, Predict All ChEMBL-Tabulated Affinities

    PubMed Central

    Cramer, Richard D.

    2015-01-01

    The possible applicability of the new template CoMFA methodology to the prediction of unknown biological affinities was explored. For twelve selected targets, all ChEMBL binding affinities were used as training and/or prediction sets, making these 3D-QSAR models the most structurally diverse and among the largest ever. For six of the targets, X-ray crystallographic structures provided the aligned templates required as input (BACE, cdk1, chk2, carbonic anhydrase-II, factor Xa, PTP1B). For all targets including the other six (hERG, cyp3A4 binding, endocrine receptor, COX2, D2, and GABAa), six modeling protocols applied to only three familiar ligands provided six alternate sets of aligned templates. The statistical qualities of the six or seven models thus resulting for each individual target were remarkably similar. Also, perhaps unexpectedly, the standard deviations of the errors of cross-validation predictions accompanying model derivations were indistinguishable from the standard deviations of the errors of truly prospective predictions. These standard deviations of prediction ranged from 0.70 to 1.14 log units and averaged 0.89 (8x in concentration units) over the twelve targets, representing an average reduction of almost 50% in uncertainty, compared to the null hypothesis of “predicting” an unknown affinity to be the average of known affinities. These errors of prediction are similar to those from Tanimoto coefficients of fragment occurrence frequencies, the predominant approach to side effect prediction, which template CoMFA can augment by identifying additional active structural classes, by improving Tanimoto-only predictions, by yielding quantitative predictions of potency, and by providing interpretable guidance for avoiding or enhancing any specific target response. PMID:26065424

  14. Bacteriophage Tailspikes and Bacterial O-Antigens as a Model System to Study Weak-Affinity Protein-Polysaccharide Interactions.

    PubMed

    Kang, Yu; Gohlke, Ulrich; Engström, Olof; Hamark, Christoffer; Scheidt, Tom; Kunstmann, Sonja; Heinemann, Udo; Widmalm, Göran; Santer, Mark; Barbirz, Stefanie

    2016-07-27

    Understanding interactions of bacterial surface polysaccharides with receptor protein scaffolds is important for the development of antibiotic therapies. The corresponding protein recognition domains frequently form low-affinity complexes with polysaccharides that are difficult to address with experimental techniques due to the conformational flexibility of the polysaccharide. In this work, we studied the tailspike protein (TSP) of the bacteriophage Sf6. Sf6TSP binds and hydrolyzes the high-rhamnose, serotype Y O-antigen polysaccharide of the Gram-negative bacterium Shigella flexneri (S. flexneri) as a first step of bacteriophage infection. Spectroscopic analyses and enzymatic cleavage assays confirmed that Sf6TSP binds long stretches of this polysaccharide. Crystal structure analysis and saturation transfer difference (STD) NMR spectroscopy using an enhanced method to interpret the data permitted the detailed description of affinity contributions and flexibility in an Sf6TSP-octasaccharide complex. Dodecasaccharide fragments corresponding to three repeating units of the O-antigen in complex with Sf6TSP were studied computationally by molecular dynamics simulations. They showed that distortion away from the low-energy solution conformation found in the octasaccharide complex is necessary for ligand binding. This is in agreement with a weak-affinity functional polysaccharide-protein contact that facilitates correct placement and thus hydrolysis of the polysaccharide close to the catalytic residues. Our simulations stress that the flexibility of glycan epitopes together with a small number of specific protein contacts provide the driving force for Sf6TSP-polysaccharide complex formation in an overall weak-affinity interaction system. PMID:27045683

  15. Ampicillin/penicillin-binding protein interactions as a model drug-target system to optimize affinity pull-down and mass spectrometric strategies for target and pathway identification.

    PubMed

    von Rechenberg, Moritz; Blake, Brian Kelly; Ho, Yew-Seng J; Zhen, Yuejun; Chepanoske, Cindy Lou; Richardson, Bonnie E; Xu, Nafei; Kery, Vladimir

    2005-05-01

    The identification and validation of the targets of active compounds identified in cell-based assays is an important step in preclinical drug development. New analytical approaches that combine drug affinity pull-down assays with mass spectrometry (MS) could lead to the identification of new targets and druggable pathways. In this work, we investigate a drug-target system consisting of ampicillin- and penicillin-binding proteins (PBPs) to evaluate and compare different amino-reactive resins for the immobilization of the affinity compound and mass spectrometric methods to identify proteins from drug affinity pull-down assays. First, ampicillin was immobilized onto various amino-reactive resins, which were compared in the ampicillin-PBP model with respect to their nonspecific binding of proteins from an Escherichia coli membrane extract. Dynal M-270 magnetic beads were chosen to further study the system as a model for capturing and identifying the targets of ampicillin, PBPs that were specifically and covalently bound to the immobilized ampicillin. The PBPs were identified, after in situ digestion of proteins bound to ampicillin directly on the beads, by using either one-dimensional (1-D) or two-dimensional (2-D) liquid chromatography (LC) separation techniques followed by tandem mass spectrometry (MS/MS) analysis. Alternatively, an elution with N-lauroylsarcosine (sarcosyl) from the ampicillin beads followed by in situ digestion and 2-D LC-MS/MS analysis identified proteins potentially interacting noncovalently with the PBPs or the ampicillin. The in situ approach required only little time, resources, and sample for the analysis. The combination of drug affinity pull-down assays with in situ digestion and 2-D LC-MS/MS analysis is a useful tool in obtaining complex information about a primary drug target as well as its protein interactors. PMID:15761956

  16. Aspects of the inverse problem for the Toda chain

    NASA Astrophysics Data System (ADS)

    Kozlowski, K. K.

    2013-12-01

    We generalize Babelon's approach to equations in dual variables so as to be able to treat new types of operators which we build out of the sub-constituents of the model's monodromy matrix. Further, we also apply Sklyanin's recent monodromy matrix identities so as to obtain equations in dual variables for yet other operators. The schemes discussed in this paper appear to be universal and thus, in principle, applicable to many models solvable through the quantum separation of variables.

  17. One- and two-dimensional Toda lattices and the Painleve property

    SciTech Connect

    Gibbon, J.D.; Tabor, M.

    1985-08-01

    The Toda lattice and the two-dimensional Toda lattice (2-DTL) are shown to possess a type of ''Painleve property'' that is based on the use of separate ''singular manifolds'' for each dependent variable. The isospectral problem for the 2-DTL found by both Mikhailov and by Fordy and Gibbons can be simply and logically derived from this analysis. Some remarks are made about the connection between our work and independent work of Kametaka and Airhault on the relationship between the Toda lattice and the second Painleve transcendent.

  18. Pyrazolo[1,5-c]quinazoline derivatives and their simplified analogues as adenosine receptor antagonists: synthesis, structure-affinity relationships and molecular modeling studies.

    PubMed

    Catarzi, Daniela; Colotta, Vittoria; Varano, Flavia; Poli, Daniela; Squarcialupi, Lucia; Filacchioni, Guido; Varani, Katia; Vincenzi, Fabrizio; Borea, Pier Andrea; Dal Ben, Diego; Lambertucci, Catia; Cristalli, Gloria

    2013-01-01

    A number of 5-oxo-pyrazolo[1,5-c]quinazolines (series B-1), bearing at position-2 the claimed (hetero)aryl moiety (compounds 1-8) but also a carboxylate group (9-14), were designed as hA(3) AR antagonists. This study produced some interesting compounds endowed with good hA(3) receptor affinity and high selectivity, being totally inactive at all the other AR subtypes. In contrast, the corresponding 5-ammino derivatives (series B-2) do not bind or bind with very low affinity at the hA(3) AR, the only exception being the 5-N-benzoyl compound 19 that shows a hA(3)K(i) value in the high μ-molar range. Evaluation of the synthetic intermediates led to the identification of some 5(3)-(2-aminophenyl)-3(5)-(hetero)arylpyrazoles 20-24 with modest affinity but high selectivity toward the hA(3) AR subtype. Molecular docking of the herein reported tricyclic and simplified derivatives was carried out to depict their hypothetical binding mode to our model of hA(3) receptor. PMID:23171656

  19. New set of symmetries and Lie algebraic structures of the Toda lattice hierarchy

    NASA Astrophysics Data System (ADS)

    Zhu, Xiao-ying; Zhang, Da-jun; Li, Zong-cheng

    2015-02-01

    By introducing the new time-dependence of the spectral parameter λ, we construct two sets of symmetries which are different from the centerless Kac-Moody-Virasoro algebras for the isospectral Toda lattice hierarchy.

  20. On the periodic Toda lattice with a self-consistent source

    NASA Astrophysics Data System (ADS)

    Babajanov, Bazar; Fečkan, Michal; Urazboev, Gayrat

    2015-05-01

    This work is devoted to the application of inverse spectral problem for integration of the periodic Toda lattice with self-consistent source. The effective method of solution of the inverse spectral problem for the discrete Hill's equation is presented.

  1. Structure-based modeling of dye-fiber affinity with SOM-4D-QSAR paradigm: application to set of anthraquinone derivatives.

    PubMed

    Bak, Andrzej; Wyszomirski, Miroslaw; Magdziarz, Tomasz; Smolinski, Adam; Polanski, Jaroslaw

    2014-01-01

    A comparative structure-affinity study of anthraquinone dyes adsorption on cellulose fibre is presented in this paper. We used receptor-dependent 4D-QSAR methods based on grid and neural (SOM) methodology coupled with IVEPLS procedure. The applied RD 4D-QSAR approach focuses mainly on the ability of mapping dye properties to verify the concept of tinctophore in dye chemistry. Moreover, the stochastic SMV procedure to investigate the predictive ability of the method for a large population of 4D-QSAR models was employed. The obtained findings were compared with the previously published RI 3D/4D-QSAR models for the corresponding anthraquinone trainings sets. The neutral (protonated) and anionic (deprotonated) forms of anthraquinone scaffold were examined in order to deal with the uncertainty of the dye ionization state. The results are comparable to both the neutral and anionic dye sets regardless of the occupancy and charge descriptors applied, respectively. It is worth noting that the SOM-4D-QSAR behaves comparably to the cubic counterpart which is observed in each training/test subset specification (4D-QSAR-Jo vs SOM- 4D-QSARo and 4D-QSAR-Jq vs SOM-4D-QSARq). Additionally, an attempt was made to specify a common set of variables contributing significantly to dye-fiber binding affinity; it was simultaneously performed for some arbitrary chosen SMV models. The presented RD 4D-QSAR methodology together with IVE-PLS procedure provides a robust and predictive modeling technique, which facilitates detailed specification of the molecular motifs significantly contributing to the fiber-dye affinity. PMID:24499310

  2. Bäcklund transformation for the first flows of the relativistic Toda hierarchy and associated properties

    NASA Astrophysics Data System (ADS)

    Choudhury, Anindya Ghose

    2013-01-01

    In this communication we study a class of one parameter dependent auto-Bäcklund transformations for the first flow of the relativistic Toda lattice and also a variant of the usual Toda lattice equation. It is shown that starting from the Hamiltonian formalism such transformations are canonical in nature with a well defined generating function. The notion of spectrality is also analyzed and the separation variables are explicitly constructed.

  3. The light asymptotic limit of conformal blocks in Toda field theory

    NASA Astrophysics Data System (ADS)

    Poghosyan, Hasmik; Poghossian, Rubik; Sarkissian, Gor

    2016-05-01

    We compute the light asymptotic limit of A n-1 Toda conformal blocks by using the AGT correspondence. We show that for certain class of CFT blocks the corresponding Nekrasov partition functions in this limit are simplified drastically being represented as a sum of a restricted class of Young diagrams. In the particular case of A 2 Toda we also compute the corresponding conformal blocks using conventional CFT techniques finding a perfect agreement with the results obtained from the Nekrasov partition functions.

  4. Characterizing traveling-wave collisions in granular chains starting from integrable limits: the case of the Korteweg-de Vries equation and the Toda lattice.

    PubMed

    Shen, Y; Kevrekidis, P G; Sen, S; Hoffman, A

    2014-08-01

    Our aim in the present work is to develop approximations for the collisional dynamics of traveling waves in the context of granular chains in the presence of precompression. To that effect, we aim to quantify approximations of the relevant Hertzian FPU-type lattice through both the Korteweg-de Vries (KdV) equation and the Toda lattice. Using the availability in such settings of both one-soliton and two-soliton solutions in explicit analytical form, we initialize such coherent structures in the granular chain and observe the proximity of the resulting evolution to the underlying integrable (KdV or Toda) model. While the KdV offers the possibility to accurately capture collisions of solitary waves propagating in the same direction, the Toda lattice enables capturing both copropagating and counterpropagating soliton collisions. The error in the approximation is quantified numerically and connections to bounds established in the mathematical literature are also given. PMID:25215797

  5. Building accurate sequence-to-affinity models from high-throughput in vitro protein-DNA binding data using FeatureREDUCE.

    PubMed

    Riley, Todd R; Lazarovici, Allan; Mann, Richard S; Bussemaker, Harmen J

    2015-01-01

    Transcription factors are crucial regulators of gene expression. Accurate quantitative definition of their intrinsic DNA binding preferences is critical to understanding their biological function. High-throughput in vitro technology has recently been used to deeply probe the DNA binding specificity of hundreds of eukaryotic transcription factors, yet algorithms for analyzing such data have not yet fully matured. Here, we present a general framework (FeatureREDUCE) for building sequence-to-affinity models based on a biophysically interpretable and extensible model of protein-DNA interaction that can account for dependencies between nucleotides within the binding interface or multiple modes of binding. When training on protein binding microarray (PBM) data, we use robust regression and modeling of technology-specific biases to infer specificity models of unprecedented accuracy and precision. We provide quantitative validation of our results by comparing to gold-standard data when available. PMID:26701911

  6. Adenine Nucleotide Translocase Is Acetylated in Vivo in Human Muscle: Modeling Predicts a Decreased ADP Affinity and Altered Control of Oxidative Phosphorylation

    PubMed Central

    2015-01-01

    Proteomics techniques have revealed that lysine acetylation is abundant in mitochondrial proteins. This study was undertaken (1) to determine the relationship between mitochondrial protein acetylation and insulin sensitivity in human skeletal muscle, identifying key acetylated proteins, and (2) to use molecular modeling techniques to understand the functional consequences of acetylation of adenine nucleotide translocase 1 (ANT1), which we found to be abundantly acetylated. Eight lean and eight obese nondiabetic subjects had euglycemic clamps and muscle biopsies for isolation of mitochondrial proteins and proteomics analysis. A number of acetylated mitochondrial proteins were identified in muscle biopsies. Overall, acetylation of mitochondrial proteins was correlated with insulin action (r = 0.60; P < 0.05). Of the acetylated proteins, ANT1, which catalyzes ADP–ATP exchange across the inner mitochondrial membrane, was acetylated at lysines 10, 23, and 92. The extent of acetylation of lysine 23 decreased following exercise, depending on insulin sensitivity. Molecular dynamics modeling and ensemble docking simulations predicted the ADP binding site of ANT1 to be a pocket of positively charged residues, including lysine 23. Calculated ADP–ANT1 binding affinities were physiologically relevant and predicted substantial reductions in affinity upon acetylation of lysine 23. Insertion of these derived binding affinities as parameters into a complete mathematical description of ANT1 kinetics predicted marked reductions in adenine nucleotide flux resulting from acetylation of lysine 23. Therefore, acetylation of ANT1 could have dramatic physiological effects on ADP–ATP exchange. Dysregulation of acetylation of mitochondrial proteins such as ANT1 therefore could be related to changes in mitochondrial function that are associated with insulin resistance. PMID:24884163

  7. Molecular modeling of the surface charging of hematite. I. The calculation of proton affinities and acidities on a surface

    NASA Astrophysics Data System (ADS)

    Wasserman, Evgeny; Rustad, James R.; Felmy, Andrew R.

    1999-03-01

    Calculation of the energy of a charged defect on a surface in supercell geometry is discussed. An important example of such a calculation is evaluation of surface proton affinities and acidities, as adding or removing a proton creates a charged unit cell. Systems with periodic boundary conditions in three spatial directions and a vacuum gap between slabs are demonstrated to be inadequate for unit cells having non-zero ionic charge and uniform neutralizing background. In such a system the calculated energy diverges linearly with the thickness of the vacuum gap. A system periodic in two directions and finite in the direction perpendicular to the surface (2-D PBC) with the neutralizing background distributed as the surface charge density is free from this problem. Furthermore, the correction for the interaction of the charged defect with its own translational images is needed to speed up the convergence to the infinite dilution limit. The expression for the asymptotic correction for the energy of interaction of a charged defect with its translational images in 2-D PBC geometry has been developed in this study. The asymptotic correction is evaluated as the interaction energy of a 2-D translationally periodic array of point charges located above and below the plate of non-uniform dielectric. This is a generalization of the method of M. Leslie and M.J. Gillan [J. Phys. C, 18 (1985) 973] for the calculation of the energy of a charged defect in bulk crystals. The usefulness of this correction was demonstrated on two test cases involving the calculation of proton affinity and acidity at the (012) surface of hematite. The proposed method is likely to be important in ab initio calculations of the energy effect of the surface protonation reactions, where computational limitations dictate a small size for the unit cell.

  8. A DFT and semiempirical model-based study of opioid receptor affinity and selectivity in a group of molecules with a morphine structural core.

    PubMed

    Bruna-Larenas, Tamara; Gómez-Jeria, Juan S

    2012-01-01

    We report the results of a search for model-based relationships between mu, delta, and kappa opioid receptor binding affinity and molecular structure for a group of molecules having in common a morphine structural core. The wave functions and local reactivity indices were obtained at the ZINDO/1 and B3LYP/6-31G(∗∗) levels of theory for comparison. New developments in the expression for the drug-receptor interaction energy expression allowed several local atomic reactivity indices to be included, such as local electronic chemical potential, local hardness, and local electrophilicity. These indices, together with a new proposal for the ordering of the independent variables, were incorporated in the statistical study. We found and discussed several statistically significant relationships for mu, delta, and kappa opioid receptor binding affinity at both levels of theory. Some of the new local reactivity indices incorporated in the theory appear in several equations for the first time in the history of model-based equations. Interaction pharmacophores were generated for mu, delta, and kappa receptors. We discuss possible differences regulating binding and selectivity in opioid receptor subtypes. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process. PMID:25379287

  9. A DFT and Semiempirical Model-Based Study of Opioid Receptor Affinity and Selectivity in a Group of Molecules with a Morphine Structural Core

    PubMed Central

    Bruna-Larenas, Tamara; Gómez-Jeria, Juan S.

    2012-01-01

    We report the results of a search for model-based relationships between mu, delta, and kappa opioid receptor binding affinity and molecular structure for a group of molecules having in common a morphine structural core. The wave functions and local reactivity indices were obtained at the ZINDO/1 and B3LYP/6-31G∗∗ levels of theory for comparison. New developments in the expression for the drug-receptor interaction energy expression allowed several local atomic reactivity indices to be included, such as local electronic chemical potential, local hardness, and local electrophilicity. These indices, together with a new proposal for the ordering of the independent variables, were incorporated in the statistical study. We found and discussed several statistically significant relationships for mu, delta, and kappa opioid receptor binding affinity at both levels of theory. Some of the new local reactivity indices incorporated in the theory appear in several equations for the first time in the history of model-based equations. Interaction pharmacophores were generated for mu, delta, and kappa receptors. We discuss possible differences regulating binding and selectivity in opioid receptor subtypes. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process. PMID:25379287

  10. The sea urchin embryo as a model for mammalian developmental neurotoxicity: ontogenesis of the high-affinity choline transporter and its role in cholinergic trophic activity.

    PubMed Central

    Qiao, Dan; Nikitina, Lyudmila A; Buznikov, Gennady A; Lauder, Jean M; Seidler, Frederic J; Slotkin, Theodore A

    2003-01-01

    Embryonic development in the sea urchin requires trophic actions of the same neurotransmitters that participate in mammalian brain assembly. We evaluated the development of the high-affinity choline transporter, which controls acetylcholine synthesis. A variety of developmental neurotoxicants affect this transporter in mammalian brain. [3H]Hemicholinium-3 binding to the transporter was found in the cell membrane fraction at stages from the unfertilized egg to pluteus, with a binding affinity comparable with that seen in mammalian brain. Over the course of development, the concentration of transporter sites rose more than 3-fold, achieving concentrations comparable with those of cholinergically enriched mammalian brain regions. Dimethylaminoethanol (DMAE), a competitive inhibitor of choline transport, elicited dysmorphology beginning at the mid-blastula stage, with anomalies beginning progressively later as the concentration of DMAE was lowered. Pretreatment, cotreatment, or delayed treatment with acetylcholine or choline prevented the adverse effects of DMAE. Because acetylcholine was protective at a lower threshold, the DMAE-induced defects were most likely mediated by its effects on acetylcholine synthesis. Transient removal of the hyaline layer enabled a charged transport inhibitor, hemicholinium-3, to penetrate sufficiently to elicit similar anomalies, which were again prevented by acetylcholine or choline. These results indicate that the developing sea urchin possesses a high-affinity choline transporter analogous to that found in the mammalian brain, and, as in mammals, the functioning of this transporter plays a key role in the developmental, trophic activity of acetylcholine. The sea urchin model may thus be useful in high-throughput screening of suspected developmental neurotoxicants. PMID:14594623

  11. New method for constructing semi-invariants and integrals of the full symmetric Toda lattice

    NASA Astrophysics Data System (ADS)

    Sorin, A. S.; Chernyakov, Yu. B.

    2015-05-01

    We consider the full symmetric representation of the Lax operator matrix of the Toda lattice, which is known as the full symmetric Toda lattice. The phase space of this system is the generic orbit of the coadjoint action of the Borel subgroup B n + of SL n (ℝ). This system is integrable. We propose a new method for constructing semi-invariants and integrals of the full symmetric Toda lattice. Using only the equations of motion for the Lax eigenvector matrix, we prove the existence of the semi-invariants that are Plücker coordinates in the corresponding projective spaces. We use these semi-invariants to construct the integrals. Our new approach provides simple exact formulas for the full set of independent semi-invariants and integrals expressed in terms of the Lax matrix and also in terms of its eigenvector and eigenvalue matrices of the full symmetric Toda lattice without using the chopping and Kostant procedures. We describe the structure of the additional integrals of motion as functions on the flag space modulo the Toda flows and show how the Plücker coordinates of different projective spaces define different families of the additional integrals.

  12. Special Report: Affinity Chromatography.

    ERIC Educational Resources Information Center

    Parikh, Indu; Cuatrecasas, Pedro

    1985-01-01

    Describes the nature of affinity chromatography and its use in purifying enzymes, studying cell interactions, exploring hormone receptors, and other areas. The potential the technique may have in treating disease is also considered. (JN)

  13. AQM router design for TCP network via input constrained fuzzy control of time-delay affine Takagi-Sugeno fuzzy models

    NASA Astrophysics Data System (ADS)

    Chang, Wen-Jer; Meng, Yu-Teh; Tsai, Kuo-Hui

    2012-12-01

    In this article, Takagi-Sugeno (T-S) fuzzy control theory is proposed as a key tool to design an effective active queue management (AQM) router for the transmission control protocol (TCP) networks. The probability control of packet marking in the TCP networks is characterised by an input constrained control problem in this article. By modelling the TCP network into a time-delay affine T-S fuzzy model, an input constrained fuzzy control methodology is developed in this article to serve the AQM router design. The proposed fuzzy control approach, which is developed based on the parallel distributed compensation technique, can provide smaller probability of dropping packets than previous AQM design schemes. Lastly, a numerical simulation is provided to illustrate the usefulness and effectiveness of the proposed design approach.

  14. Structure-based model profiles affinity constant of drugs with hPEPT1 for rapid virtual screening of hPEPT1's substrate.

    PubMed

    Sun, L; Meng, S

    2016-08-01

    The human proton-coupled peptide transporter (hPEPT1) with broad substrates is an important route for improving the pharmacokinetic performance of drugs. Thus, it is essential to predict the affinity constant between drug molecule and hPEPT1 for rapid virtual screening of hPEPT1's substrate during lead optimization, candidate selection and hPEPT1 prodrug design. Here, a structure-based in silico model for 114 compounds was constructed based on eight structural parameters. This model was built by the multiple linear regression method and satisfied all the prerequisites of the regression models. For the entire data set, the r(2) and adjusted r(2) values were 0.74 and 0.72, respectively. Then, this model was used to perform substrate/non-substrate classification. For 29 drugs from DrugBank database, all were correctly classified as substrates of hPEPT1. This model was also used to perform substrate/non-substrate classification for 18 drugs and their prodrugs; this QSAR model also can distinguish between the substrate and non-substrate. In conclusion, the QSAR model in this paper was validated by a large external data set, and all results indicated that the developed model was robust, stable, and can be used for rapid virtual screening of hPEPT1's substrate in the early stage of drug discovery. PMID:27586363

  15. Constrained lattice-field hierarchies and Toda system with Block symmetry

    NASA Astrophysics Data System (ADS)

    Li, Chuanzhong

    2016-03-01

    In this paper, we construct the additional W-symmetry and ghost symmetry of two-lattice field integrable hierarchies. Using the symmetry constraint, we construct constrained two-lattice integrable systems which contain several new integrable difference equations. Under a further reduction, the constrained two-lattice integrable systems can be combined into one single integrable system, namely the well-known one-dimensional original Toda hierarchy. We prove that the one-dimensional original Toda hierarchy has a nice Block Lie symmetry.

  16. Quasi-periodic solutions to the hierarchy of four-component Toda lattices

    NASA Astrophysics Data System (ADS)

    Wei, Jiao; Geng, Xianguo; Zeng, Xin

    2016-08-01

    Starting from a discrete 3×3 matrix spectral problem, the hierarchy of four-component Toda lattices is derived by using the stationary discrete zero-curvature equation. Resorting to the characteristic polynomial of the Lax matrix for the hierarchy, we introduce a trigonal curve Km-2 of genus m - 2 and present the related Baker-Akhiezer function and meromorphic function on it. Asymptotic expansions for the Baker-Akhiezer function and meromorphic function are given near three infinite points on the trigonal curve, from which explicit quasi-periodic solutions for the hierarchy of four-component Toda lattices are obtained in terms of the Riemann theta function.

  17. Kernel functions and Baecklund transformations for relativistic Calogero-Moser and Toda systems

    SciTech Connect

    Hallnaes, Martin; Ruijsenaars, Simon

    2012-12-15

    We obtain kernel functions associated with the quantum relativistic Toda systems, both for the periodic version and for the nonperiodic version with its dual. This involves taking limits of previously known results concerning kernel functions for the elliptic and hyperbolic relativistic Calogero-Moser systems. We show that the special kernel functions at issue admit a limit that yields generating functions of Baecklund transformations for the classical relativistic Calogero-Moser and Toda systems. We also obtain the nonrelativistic counterparts of our results, which tie in with previous results in the literature.

  18. Ionization Energies, Electron Affinities, and Polarization Energies of Organic Molecular Crystals: Quantitative Estimations from a Polarizable Continuum Model (PCM)-Tuned Range-Separated Density Functional Approach.

    PubMed

    Sun, Haitao; Ryno, Sean; Zhong, Cheng; Ravva, Mahesh Kumar; Sun, Zhenrong; Körzdörfer, Thomas; Brédas, Jean-Luc

    2016-06-14

    We propose a new methodology for the first-principles description of the electronic properties relevant for charge transport in organic molecular crystals. This methodology, which is based on the combination of a nonempirical, optimally tuned range-separated hybrid functional with the polarizable continuum model, is applied to a series of eight representative molecular semiconductor crystals. We show that it provides ionization energies, electron affinities, and transport gaps in very good agreement with experimental values, as well as with the results of many-body perturbation theory within the GW approximation at a fraction of the computational costs. Hence, this approach represents an easily applicable and computationally efficient tool to estimate the gas-to-crystal phase shifts of the frontier-orbital quasiparticle energies in organic electronic materials. PMID:27183355

  19. The generalized Toda lattices and the Whitham averaged system for the defocusing nonlinear Schrodinger equation

    NASA Astrophysics Data System (ADS)

    Ye, Jian

    In Part I, we first study an iso-spectral deformation of general matrix which is a natural generalization of the nonperiodic Toda lattice equation. This deformation is equivalent to the Cholesky flow. We prove the integrability of the deformation, and give an explicit formula for the solution to the initial value problem. The formula is obtained by generalizing the orthogonalization procedure of Szego. Using the formula, the solution to the LU factorization can be constructed explicitly. Based on the root spaces for simple Lie algebras, we consider several reductions of the equation. This leads to generalized Toda equations related to other classical semi-simple Lie algebra which include the integrable systems studied by Bogoyavlensky and Kostant. We show these systems can be solved explicitly in a unified way. Based on the explicit solutions, we then consider the iso-spectral real manifolds of tridiagonal Hessenberg matrices with distinct real eigenvalues. The manifolds are described by the iso-spectral flows of indefinite Toda lattice equations introduced by Kodama and Ye. These Toda lattices consist of 2N-1 different systems with hamiltonians H = [[1]/over[2

  20. Building accurate sequence-to-affinity models from high-throughput in vitro protein-DNA binding data using FeatureREDUCE

    PubMed Central

    Riley, Todd R; Lazarovici, Allan; Mann, Richard S; Bussemaker, Harmen J

    2015-01-01

    Transcription factors are crucial regulators of gene expression. Accurate quantitative definition of their intrinsic DNA binding preferences is critical to understanding their biological function. High-throughput in vitro technology has recently been used to deeply probe the DNA binding specificity of hundreds of eukaryotic transcription factors, yet algorithms for analyzing such data have not yet fully matured. Here, we present a general framework (FeatureREDUCE) for building sequence-to-affinity models based on a biophysically interpretable and extensible model of protein-DNA interaction that can account for dependencies between nucleotides within the binding interface or multiple modes of binding. When training on protein binding microarray (PBM) data, we use robust regression and modeling of technology-specific biases to infer specificity models of unprecedented accuracy and precision. We provide quantitative validation of our results by comparing to gold-standard data when available. DOI: http://dx.doi.org/10.7554/eLife.06397.001 PMID:26701911

  1. An explicitly solvated full atomistic model of the cardiac thin filament and application on the calcium binding affinity effects from familial hypertrophic cardiomyopathy linked mutations

    NASA Astrophysics Data System (ADS)

    Williams, Michael; Schwartz, Steven

    2015-03-01

    The previous version of our cardiac thin filament (CTF) model consisted of the troponin complex (cTn), two coiled-coil dimers of tropomyosin (Tm), and 29 actin units. We now present the newest revision of the model to include explicit solvation. The model was developed to continue our study of genetic mutations in the CTF proteins which are linked to familial hypertrophic cardiomyopathies. Binding of calcium to the cTnC subunit causes subtle conformational changes to propagate through the cTnC to the cTnI subunit which then detaches from actin. Conformational changes propagate through to the cTnT subunit, which allows Tm to move into the open position along actin, leading to muscle contraction. Calcium disassociation allows for the reverse to occur, which results in muscle relaxation. The inclusion of explicit TIP3 water solvation allows for the model to get better individual local solvent to protein interactions; which are important when observing the N-lobe calcium binding pocket of the cTnC. We are able to compare in silica and in vitro experimental results to better understand the physiological effects from mutants, such as the R92L/W and F110V/I of the cTnT, on the calcium binding affinity compared to the wild type.

  2. Investigation of the Chromosome Regions with Significant Affinity for the Nuclear Envelope in Fruit Fly – A Model Based Approach

    PubMed Central

    Kinney, Nicholas Allen; Sharakhov, Igor V.; Onufriev, Alexey V.

    2014-01-01

    Three dimensional nuclear architecture is important for genome function, but is still poorly understood. In particular, little is known about the role of the “boundary conditions” – points of attachment between chromosomes and the nuclear envelope. We describe a method for modeling the 3D organization of the interphase nucleus, and its application to analysis of chromosome-nuclear envelope (Chr-NE) attachments of polytene (giant) chromosomes in Drosophila melanogaster salivary glands. The model represents chromosomes as self-avoiding polymer chains confined within the nucleus; parameters of the model are taken directly from experiment, no fitting parameters are introduced. Methods are developed to objectively quantify chromosome territories and intertwining, which are discussed in the context of corresponding experimental observations. In particular, a mathematically rigorous definition of a territory based on convex hull is proposed. The self-avoiding polymer model is used to re-analyze previous experimental data; the analysis suggests 33 additional Chr-NE attachments in addition to the 15 already explored Chr-NE attachments. Most of these new Chr-NE attachments correspond to intercalary heterochromatin – gene poor, dark staining, late replicating regions of the genome; however, three correspond to euchromatin – gene rich, light staining, early replicating regions of the genome. The analysis also suggests 5 regions of anti-contact, characterized by aversion for the NE, only two of these correspond to euchromatin. This composition of chromatin suggests that heterochromatin may not be necessary or sufficient for the formation of a Chr-NE attachment. To the extent that the proposed model represents reality, the confinement of the polytene chromosomes in a spherical nucleus alone does not favor the positioning of specific chromosome regions at the NE as seen in experiment; consequently, the 15 experimentally known Chr-NE attachment positions do not appear to

  3. Dannie Heineman Prize for Mathematical Physics Prize Lecture: Correlation Functions in Integrable Models II: The Role of Quantum Affine Symmetry

    NASA Astrophysics Data System (ADS)

    Jimbo, Michio

    2013-03-01

    Since the beginning of 1980s, hidden infinite dimensional symmetries have emerged as the origin of integrability: first in soliton theory and then in conformal field theory. Quest for symmetries in quantum integrable models has led to the discovery of quantum groups. On one hand this opened up rapid mathematical developments in representation theory, combinatorics and other fields. On the other hand it has advanced understanding of correlation functions of lattice models, leading to multiple integral formulas in integrable spin chains. We shall review these developments which continue up to the present time.

  4. Solutions of the sDiff(2)Toda equation with SU(2) symmetry

    NASA Astrophysics Data System (ADS)

    Finley, Daniel; McIver, John K.

    2010-07-01

    We present the general solution to the Plebański equation for an \\bm{\\mathfrak{h}} space that admits Killing vectors for an entire SU(2) of symmetries, which is therefore also the general solution of the sDiff(2)Toda equation that allows these symmetries. Desiring these solutions as a bridge toward the future for yet more general solutions of the sDiff(2)Toda equation, we generalize the earlier work of Olivier, on the Atiyah-Hitchin metric, and re-formulate work of Babich and Korotkin, and Tod, on the Bianchi IX approach to a metric with an SU(2) of symmetries. We also give careful delineations of the conformal transformations required to ensure that a metric of Bianchi IX type has a zero Ricci tensor, so that it is a self-dual, vacuum solution of the complex-valued version of Einstein's equations, as appropriate for the original Plebański equation.

  5. On the evolution of scattering data under perturbations of the Toda lattice

    NASA Astrophysics Data System (ADS)

    Bilman, D.; Nenciu, I.

    2016-09-01

    We present the results of an analytical and numerical study of the long-time behavior for certain Fermi-Pasta-Ulam (FPU) lattices viewed as perturbations of the completely integrable Toda lattice. Our main tools are the direct and inverse scattering transforms for doubly-infinite Jacobi matrices, which are well-known to linearize the Toda flow. We focus in particular on the evolution of the associated scattering data under the perturbed vs. the unperturbed equations. We find that the eigenvalues present initially in the scattering data converge to new, slightly perturbed eigenvalues under the perturbed dynamics of the lattice equation. To these eigenvalues correspond solitary waves that emerge from the solitons in the initial data. We also find that new eigenvalues emerge from the continuous spectrum as the lattice system is let to evolve under the perturbed dynamics.

  6. The Full Kostant-Toda Hierarchy on the Positive Flag Variety

    NASA Astrophysics Data System (ADS)

    Kodama, Yuji; Williams, Lauren

    2015-04-01

    We study some geometric and combinatorial aspects of the solution to the full Kostant-Toda (f-KT) hierarchy, when the initial data is given by an arbitrary point on the totally non-negative (tnn) flag variety of . The f-KT flows on the tnn flag variety are complete, and we show that their asymptotics are completely determined by the cell decomposition of the tnn flag variety given by Rietsch (Total positivity and real flag varieties. Ph.D. Thesis, Massachusetts Institute of Technology, Cambridge, 1998). Our results represent the first results on the asymptotics of the f-KT hierarchy (and even the f-KT lattice); moreover, our results are not confined to the generic flow, but cover non-generic flows as well. We define the f-KT flow on the weight space via the moment map, and show that the closure of each f-KT flow forms an interesting convex polytope which we call a Bruhat interval polytope. In particular, the Bruhat interval polytope for the generic flow is the permutohedron of the symmetric group . We also prove analogous results for the full symmetric Toda hierarchy, by mapping our f-KT solutions to those of the full symmetric Toda hierarchy. In the appendix we show that Bruhat interval polytopes are generalized permutohedra, in the sense of Postnikov (Int. Math. Res. Not. IMRN (6):1026-1106, 2009).

  7. Effects of Water Models on Binding Affinity: Evidence from All-Atom Simulation of Binding of Tamiflu to A/H5N1 Neuraminidase

    PubMed Central

    Nguyen, Trang Truc; Viet, Man Hoang

    2014-01-01

    The influence of water models SPC, SPC/E, TIP3P, and TIP4P on ligand binding affinity is examined by calculating the binding free energy ΔGbind of oseltamivir carboxylate (Tamiflu) to the wild type of glycoprotein neuraminidase from the pandemic A/H5N1 virus. ΔGbind is estimated by the Molecular Mechanic-Poisson Boltzmann Surface Area method and all-atom simulations with different combinations of these aqueous models and four force fields AMBER99SB, CHARMM27, GROMOS96 43a1, and OPLS-AA/L. It is shown that there is no correlation between the binding free energy and the water density in the binding pocket in CHARMM. However, for three remaining force fields ΔGbind decays with increase of water density. SPC/E provides the lowest binding free energy for any force field, while the water effect is the most pronounced in CHARMM. In agreement with the popular GROMACS recommendation, the binding score obtained by combinations of AMBER-TIP3P, OPLS-TIP4P, and GROMOS-SPC is the most relevant to the experiments. For wild-type neuraminidase we have found that SPC is more suitable for CHARMM than TIP3P recommended by GROMACS for studying ligand binding. However, our study for three of its mutants reveals that TIP3P is presumably the best choice for CHARMM. PMID:24672329

  8. Modeling Condensation, Hydro- and Pepto-affinity of Surfaces in Medical Implant Devices and Surgical Lenses: Effect of Blood Proteins

    NASA Astrophysics Data System (ADS)

    Bennett-Kennett, Ross; Herbots, Nicole; Murphy, Ashlee; Sell, David; Kutz, Tyler; Benitez, Sophia; Acharya, Ajjya; Hughes, Brett; Watson, Clarizza; Culbertson, Eric; Sell, Clive; Kwong, H.

    2012-10-01

    Surgical lenses in laparoscopes and arthroscopes ``fog'' during surgery. Fogging increases by up to 40% surgery duration, infection rates, and scarring due to exposure from repeated scopes withdrawal for cleaning. Modeling nucleation on surfaces shows that 2-D layer-by-layer condensation maintains transparency while 3-D droplets refract at gas/fluid interfaces leading to opacity or ``fogging.'' This ProteinKnoxmodel for lenses made from bio-compatible polymers, and silica led us to a nano-scale molecular mesh applied as a bio-identical emulsion. ProteinKnox[1-5] meets a 100% success rate in eliminating fogging for up to 240 minutes over 300 experiments. Twenty surgical trials in the OR yield a success rate of 90%, with loss of vision due to the presence of blood or blood proteins, not fogging. We studied the common blood protein, heparin, which prevents coagulation, with the ProteinKnoxmodel. Heparin behaves like H2O on hydrophobic surfaces. It does not prevent fogging nor interferes with 2-D condensatio. Next, we investigated fibrinogen as agonist agent because it causes coagulation. Fibrinogen applied to various surfaces in emulsions prepared in accordance with the ProteinKnoxmodel can prevent not only

  9. Combination of crossflow ultrafiltration, monolithic affinity filtration, and quantitative reverse transcriptase PCR for rapid concentration and quantification of model viruses in water.

    PubMed

    Pei, Lu; Rieger, Martin; Lengger, Sandra; Ott, Sonja; Zawadsky, Claudia; Hartmann, Nils Marten; Selinka, Hans-Christoph; Tiehm, Andreas; Niessner, Reinhard; Seidel, Michael

    2012-09-18

    We present a rapid and effective adsorption-elution method based on monolithic affinity filtration (MAF) for the concentration and purification of waterborne viruses. The MAF column consists of a hydrolyzed macroporous epoxy-based polymer. High recoveries were achieved by columns for the bacterial virus (bacteriophage) MS2 110 (±19)%, as model organism, as well as for human adenoviruses 42.4 (±3.4)% and murine noroviruses 42.6 (±1.9)%. This new concentration and purification method was combined with crossflow ultrafiltration (CUF). Because of the adsorption of the examined viruses to the macroporous surface of the MAF column at pH 3, concentrated matrix components by CUF can be removed. Bacteriophages MS2 were spiked in tap water and concentrated with the new CUF-MAF concentration method by a volumetric factor of 10(4) within 33 min. Furthermore, the detection limit for quantification of bacteriophage MS2 by quantitative reverse transcriptase PCR (qRT-PCR) could be improved from 79.47 to 0.0056 GU mL(-1) by a factor of 1.4 × 10(4). In a first study, we have shown that this method could also be applied for river water containing naturally MS2 and MS2-like phages. PMID:22917471

  10. Biomimetic design of platelet adhesion inhibitors to block integrin α2β1-collagen interactions: I. Construction of an affinity binding model.

    PubMed

    Zhang, Lin; Sun, Yan

    2014-04-29

    Platelet adhesion on a collagen surface through integrin α2β1 has been proven to be significant for the formation of arterial thrombus. However, the molecular determinants mediating the integrin-collagen complex remain unclear. In the present study, the dynamics of integrin-collagen binding and molecular interactions were investigated using molecular dynamics (MD) simulations and molecular mechanics-Poisson-Boltzmann surface area (MM-PBSA) analysis. Hydrophobic interaction is identified as the major driving force for the formation of the integrin-collagen complex. On the basis of the MD simulation and MM-PBSA results, an affinity binding model (ABM) of integrin for collagen is constructed; it is composed of five residues, including Y157, N154, S155, R288, and L220. The ABM has been proven to capture the major binding motif contributing 84.8% of the total binding free energy. On the basis of the ABM, we expect to establish a biomimetic design strategy of platelet adhesion inhibitors, which would be beneficial for the development of potent peptide-based drugs for thrombotic diseases. PMID:24697616

  11. Pulmonary Administration of GW0742, a High-Affinity Peroxisome Proliferator-Activated Receptor Agonist, Repairs Collapsed Alveoli in an Elastase-Induced Mouse Model of Emphysema.

    PubMed

    Ozawa, Chihiro; Horiguchi, Michiko; Akita, Tomomi; Oiso, Yuki; Abe, Kaori; Motomura, Tomoki; Yamashita, Chikamasa

    2016-01-01

    Pulmonary emphysema is a disease in which lung alveoli are irreversibly damaged, thus compromising lung function. Our previous study revealed that all-trans-retinoic acid (ATRA) induces the differentiation of human lung alveolar epithelial type 2 progenitor cells and repairs the alveoli of emphysema model mice. ATRA also reportedly has the ability to activate peroxisome proliferator-activated receptor (PPAR) β/δ. A selective PPARβ/δ ligand has been reported to induce the differentiation of human keratinocytes during wound repair. Here, we demonstrate that treatment using a high-affinity PPARβ/δ agonist, GW0742, reverses the lung tissue damage induced by elastase in emphysema-model mice and improves respiratory function. Mice treated with elastase, which collapsed their alveoli, were then treated with either 10% dimethyl sulfoxide (DMSO) in saline (control group) or GW0742 (1.0 mg/kg twice a week) by pulmonary administration. Treatment with GW0742 for 2 weeks increased the in vivo expression of surfactant proteins A and D, which are known alveolar type II epithelial cell markers. GW0742 treatment also shortened the average distance between alveolar walls in the lungs of emphysema model mice, compared with a control group treated with 10% DMSO in saline. Treatment with GW0742 for 3 weeks also improved tissue elastance (cm H2O/mL), as well as the ratio of the forced expiratory volume in the first 0.05 s to the forced vital capacity (FEV 0.05/FVC). In each of these experiments, GW0742 treatment reversed the damage caused by elastase. In conclusion, PPARβ/δ agonists are potential therapeutic agents for pulmonary emphysema. PMID:27150147

  12. Local Affinity Release.

    PubMed

    Delplace, Vianney; Obermeyer, Jaclyn; Shoichet, Molly S

    2016-07-26

    The use of hydrogels for therapeutic delivery is a burgeoning area of investigation. These water-swollen polymer matrices are ideal platforms for localized drug delivery that can be further combined with specific ligands or nanotechnologies to advance the controlled release of small-molecule drugs and proteins. Due to the advantage of hydrophobic, electrostatic, or specific extracellular matrix interactions, affinity-based strategies can overcome burst release and challenges associated with encapsulation. Future studies will provide innovative binding tools, truly stimuli-responsive systems, and original combinations of emerging technologies to control the release of therapeutics spatially and temporally. Local drug delivery can be achieved by directly injecting a therapeutic to its site of action and is advantageous because off-target effects associated with systemic delivery can be minimized. For prolonged benefit, a vehicle that provides sustained drug release is required. Hydrogels are versatile platforms for localized drug release, owing to the large library of biocompatible building blocks from which they can be formed. Injectable hydrogel formulations that gel quickly in situ and provide sustained release of therapeutics are particularly advantageous to minimize invasiveness. The incorporation of polymers, ligands or nanoparticles that have an affinity for the therapeutic of interest improve control over the release of small-molecule drugs and proteins from hydrogels, enabling spatial and temporal control over the delivery. Such affinity-based strategies can overcome drug burst release and challenges associated with protein instability, allowing more effective therapeutic molecule delivery for a range of applications from therapeutic contact lenses to ischemic tissue regeneration. PMID:27403513

  13. Identification, Modeling and Ligand Affinity of Early Deuterostome CYP51s, and Functional Characterization of Recombinant Zebrafish Sterol 14α-Demethylase

    PubMed Central

    Morrison, Ann Michelle Stanley; Goldstone, Jared V.; Lamb, David C.; Kubota, Akira; Lemaire, Benjamin; Stegeman, John. J.

    2014-01-01

    Background Sterol 14α-demethylase (cytochrome P450 51, CYP51, P45014DM) is a microsomal enzyme that in eukaryotes catalyzes formation of sterols essential for cell membrane function and as precursors in biosynthesis of steroid hormones. Functional properties of CYP51s are unknown in non-mammalian deuterostomes. Methods PCR-cloning and sequencing and computational analyses (homology modeling and docking) addressed CYP51 in zebrafish Danio rerio, the reef fish sergeant major Abudefduf saxatilis, and the sea urchin Strongylocentrotus purpuratus. Following N-terminal amino acid modification, zebrafish CYP51 was expressed in Escherichia coli, and lanosterol 14α-demethylase activity and azole inhibition of CYP51 activity were characterized using GC/MS. Results Molecular phylogeny positioned S. purpuratus CYP51 at the base of the deuterostome clade. In zebrafish, CYP51 is expressed in all organs examined, most strongly in intestine. The recombinant protein bound lanosterol and catalyzed 14α-demethylase activity, at 3.2 nmol/min/nmol CYP51. The binding of azoles to zebrafish CYP51 gave KS (dissociation constant) values of 0.26 μM for ketoconazole and 0.64 μM for propiconazole. Displacement of carbon monoxide also indicated zebrafish CYP51 has greater affinity for ketoconazole. Docking to homology models showed that lanosterol docks in fish and sea urchin CYP51s with an orientation essentially the same as in mammalian CYP51. Docking of ketoconazole indicates it would inhibit fish and sea urchin CYP51s. Conclusions Biochemical and computational analyses are consistent with lanosterol being a substrate for early deuterostome CYP51s. General Significance The results expand the phylogenetic view of animal CYP51, with evolutionary, environmental and therapeutic implications. PMID:24361620

  14. Morphometric affinities of gigantopithecus.

    PubMed

    Gelvin, B R

    1980-11-01

    Multivariate analyses, supplemented by univariate statistical methods, of measurements from mandibular tooth crown dimensions and the mandible of Gigantopithecus blacki, G. bilaspurensis, Plio-Plelstocene hominids, Homo erectus, and seven Neogene ape species from the genera Proconsul, Sivapithecus, Ouranopithecus, and Dryopithecus were used to assess the morphometric affinities of Gigantopithecus. The results show that Gigantopithecus displays affinities to Ouranopithecus and to the hominids, particularly the Plio-Plelstocene hominids, rather than to the apes. Ouranopithecus demonstrated dental resemblances to G. bilaspurensis and the Plio-Pleistocene hominids but mandibular similarities to the apes. Results of analyses of tooth and mandibular shape indices, combined with multivariate distance and temporal relationships, suggest that Ouranopithecus is a more likely candidate for Gigantopithecus ancestry than is Silvapithecus indicus. Shape and allometric differences between G. bilaspurensis and the robust australopithecines weaken the argument for an ancestral-descendant relationship between these groups. The results support the hypothesis that Gigantopithecus is an extinct side branch of the Hominidae. PMID:7468790

  15. Correlation functions with fusion-channel multiplicity in {mathcal{W}}_3 Toda field theory

    NASA Astrophysics Data System (ADS)

    Belavin, Vladimir; Estienne, Benoit; Foda, Omar; Santachiara, Raoul

    2016-06-01

    Current studies of {mathcal{W}}_N Toda field theory focus on correlation functions such that the {mathcal{W}}_N highest-weight representations in the fusion channels are multiplicity-free. In this work, we study {mathcal{W}}_3 Toda 4-point functions with multiplicity in the fusion channel. The conformal blocks of these 4-point functions involve matrix elements of a fully-degenerate primary field with a highest-weight in the adjoint representation of mathfrak{s}{mathfrak{l}}_3 , and a fully-degenerate primary field with a highest-weight in the fundamental representation of mathfrak{s}{mathfrak{l}}_3 . We show that, when the fusion rules do not involve multiplicities, the matrix elements of the fully-degenerate adjoint field, between two arbitrary descendant states, can be computed explicitly, on equal footing with the matrix elements of the semi-degenerate fundamental field. Using null-state conditions, we obtain a fourth-order Fuchsian differential equation for the conformal blocks. Using Okubo theory, we show that, due to the presence of multiplicities, this differential equation belongs to a class of Fuchsian equations that is different from those that have appeared so far in {mathcal{W}}_N theories. We solve this equation, compute its monodromy group, and construct the monodromy-invariant correlation functions. This computation shows in detail how the ambiguities that are caused by the presence of multiplicities are fixed by requiring monodromy-invariance.

  16. Asymptotics of a Class of Solutions to the Cylindrical Toda Equations

    NASA Astrophysics Data System (ADS)

    Tracy, Craig A.; Widom, Harold

    The small t asymptotics of a class of solutions to the 2D cylindrical Toda equations is computed. The solutions, , have the representation where Kk$ are integral operators. This class includes the n-periodic cylindrical Toda equations. For n=2 our results reduce to the previously computed asymptotics of the 2D radial sinh-Gordon equation and for n=3 (and with an additional symmetry constraint) they reduce to earlier results for the radial Bullough-Dodd equation. Both of these special cases are examples of Painlevé III and have arisen in various applications. The asymptotics of are derived by computing the small t asymptotics where explicit formulas are given for the quantities ak and bk. The method consists of showing that the resolvent operator of Kk has an approximation in terms of resolvents of certain Wiener-Hopf operators, for which there are explicit integral formulas.

  17. A High-Affinity Native Human Antibody Disrupts Biofilm from Staphylococcus aureus Bacteria and Potentiates Antibiotic Efficacy in a Mouse Implant Infection Model.

    PubMed

    Estellés, Angeles; Woischnig, Anne-Kathrin; Liu, Keyi; Stephenson, Robert; Lomongsod, Evelene; Nguyen, Da; Zhang, Jianzhong; Heidecker, Manfred; Yang, Yifan; Simon, Reyna J; Tenorio, Edgar; Ellsworth, Stote; Leighton, Anton; Ryser, Stefan; Gremmelmaier, Nina Khanna; Kauvar, Lawrence M

    2016-04-01

    Many serious bacterial infections are difficult to treat due to biofilm formation, which provides physical protection and induces a sessile phenotype refractory to antibiotic treatment compared to the planktonic state. A key structural component of biofilm is extracellular DNA, which is held in place by secreted bacterial proteins from the DNABII family: integration host factor (IHF) and histone-like (HU) proteins. A native human monoclonal antibody, TRL1068, has been discovered using single B-lymphocyte screening technology. It has low-picomolar affinity against DNABII homologs from important Gram-positive and Gram-negative bacterial pathogens. The disruption of established biofilm was observedin vitroat an antibody concentration of 1.2 μg/ml over 12 h. The effect of TRL1068in vivowas evaluated in a murine tissue cage infection model in which a biofilm is formed by infection with methicillin-resistantStaphylococcus aureus(MRSA; ATCC 43300). Treatment of the established biofilm by combination therapy of TRL1068 (15 mg/kg of body weight, intraperitoneal [i.p.] administration) with daptomycin (50 mg/kg, i.p.) significantly reduced adherent bacterial count compared to that after daptomycin treatment alone, accompanied by significant reduction in planktonic bacterial numbers. The quantification of TRL1068 in sample matrices showed substantial penetration of TRL1068 from serum into the cage interior. TRL1068 is a clinical candidate for combination treatment with standard-of-care antibiotics to overcome the drug-refractory state associated with biofilm formation, with potential utility for a broad spectrum of difficult-to-treat bacterial infections. PMID:26833157

  18. Importance of protein flexibility in ranking inhibitor affinities: modeling the binding mechanisms of piperidine carboxamides as Type I1/2 ALK inhibitors.

    PubMed

    Kong, Xiaotian; Pan, Peichen; Li, Dan; Tian, Sheng; Li, Youyong; Hou, Tingjun

    2015-02-28

    Anaplastic lymphoma kinase (ALK) has gained increased attention as an attractive therapeutic target for the treatment of various cancers, especially non-small-cell lung cancer (NSCLC). Recently, piperidine carboxamides were reported as Type I1/2 inhibitors of ALK, which occupy both the ATP binding site and the back ATP hydrophobic cavity in DFG-in conformation. Due to the dynamic behavior of ALK in the binding of Type I1/2 inhibitors, the accurate predictions of the binding structures and relative binding potencies of these inhibitors are quite challenging. In this study, different modeling techniques, including molecular docking, ensemble docking based on multiple receptor conformations, molecular dynamics simulations and free energy calculations, were utilized to explore the binding mechanisms of piperidine carboxamides. Our predictions show that the conventional docking protocols are not sufficient to predict the relative binding potencies of the studied inhibitors with high accuracy, but incorporating protein flexibility before or after docking is quite effective to improve the prediction accuracy. Notably, the binding free energies predicted by MM/GBSA or MM/PBSA based on the MD simulations for the docked poses give the highest correlation with the experimental data, highlighting the importance of the inclusion of receptor flexibility for the accurate predictions of the binding potencies for Type I1/2 inhibitors of ALK. Furthermore, the comprehensive analysis of several pairs of representative inhibitors demonstrates the importance of hydrophobic interactions in improving the binding affinities of the inhibitors with the hot-spot residues surrounding the binding pocket. This work is expected to provide valuable clues for further rational design of novel and potent Type I1/2 ALK inhibitors. PMID:25644934

  19. Adjoint affine fusion and tadpoles

    NASA Astrophysics Data System (ADS)

    Urichuk, Andrew; Walton, Mark A.

    2016-06-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  20. Mechanism-Based Competitive Binding Model to Investigate the Effect of Neonatal Fc Receptor Binding Affinity on the Pharmacokinetic of Humanized Anti-VEGF Monoclonal IgG1 Antibody in Cynomolgus Monkey.

    PubMed

    Ng, Chee M; Fielder, Paul J; Jin, Jin; Deng, Rong

    2016-07-01

    The quantitative relationship between neonatal Fc receptor (FcRn) binding affinity at both acidic and physiological pH and the pharmacokinetics of protein engineered FcRn IgG1 variants has not yet been reported. Our objective was to develop a quantitatively mechanism-based competitive binding model to describe the effects of FcRn binding affinity at acidic and physiological pH on the pharmacokinetics of anti-VEGF IgG1 antibodies when both endogenous and exogenous antibodies are competing for the same FcRn. Pharmacokinetic (PK) and FcRn binding data from five Fc variants of humanized anti-VEGF IgG1 monoclonal antibodies with wide range of FcRn binding affinity were used for the analysis. Sixty-seven anti-VEGF IgG1 antibody-treated animals and 25 control animals with simulated endogenous IgG levels were used to develop the final model. A hybrid iterative two stages and Monte Carlo parametric expectation-maximization method was used to obtain the final model parameters estimates. The final model well described the observed PK data. Quantitative FcRn binding affinity-pharmacokinetics relationships was constructed to provide important biological insights in better understanding of the FcRn binding effect on pharmacokinetics of anti-VEGF IgG1 antibodies in cynomolgus monkeys and served as an important model-based drug discovery platform to guide the design and development of the future generation of anti-VEGF or other therapeutic IgG1 antibodies. PMID:27075465

  1. Coordination of two high-affinity hexamer peptides to copper(II) and palladium(II) models of the peptide-metal chelation site on IMAC resins

    SciTech Connect

    Chen, Y.; Pasquinelli, R.; Ataai, M.; Koepsel, R.R.; Kortes, R.A.; Shepherd, R.E.

    2000-03-20

    The coordination of peptides Ser-Pro-His-His-Gly-Gly (SPHHGG) and (His){sub 6} (HHHHHH) to [Pd{sup II}(mida)(D{sub 2}O)] (mida{sup 2{minus}} = N-methyliminodiacetate) was studied by {sup 1}H NMR as model reactions for Cu{sup II}(iminodiacetate)-immobilized metal affinity chromatography (IMAC) sites. This is the first direct physical description of peptide coordination for IMAC. A three-site coordination is observed which involves the first, third, and fourth residues along the peptide chain. The presence of proline in position 2 of SPHHGG achieves the best molecular mechanics and bonding angles in the coordinated peptide and enhances the interaction of the serine amino nitrogen. Histidine coordination of H{sub 1}, H{sub 3}, and H{sub 4} of (His){sub 6} and H{sub 3} and H{sub 4} of SPHHGG was detected by {sup 1}H NMR contact shifts and H/D exchange of histidyl protons. The EPR spectra of SPHHGG and HHHHHH attached to the [Cu{sup II}(mida)] unit were obtained for additional modeling of IMAC sites. EPR parameters of the parent [Cu(mida)(H{sub 2}O){sub 2}] complex are representative: g{sub zz} = 2.31; g{sub yy} = 2.086; g{sub xx} = 2.053; A{sub {vert_bar}{vert_bar}} = 161 G; A{sub N} = 19G (three line, one N coupling). Increased rhombic distortion is detected relative to the starting aqua complex in the order of [Cu(mida)L] for distortion of HHHHHH > SPHHGG > (H{sub 2}O){sub 2}. The lowering of symmetry is also seen in the decrease in the N-shf coupling, presumably to the imino nitrogen of mida{sup 2{minus}} in the order 19 G (H{sub 2}O), 16 G (SPHHGG) and 11 G (HHHHHH). Visible spectra of the [Cu(mida)(SPHHGG)] and [Cu(mida)(HHHHHH)] as a function of pH indicate coordination of one histidyl donor at ca. 4.5, two in the range of pH 5--7, and two chelate ring attachments involving the terminal amino donor for SPHHGG or another histidyl donor of HHHHHH in the pH domain of 7--8 in agreement with the [Pd{sup II}(mida)L] derivatives which form the two

  2. FURTHER MAPPING OF THE NATALITY CHRONOME IN TODA CITY (JAPAN) MATERNITY HOSPITAL

    PubMed Central

    YAMANAKA, T.; CORNÉLISSEN, G.; KAZUMA, M.; KAZUMA, N.; MURAKAMI, S.; OTSUKA, K.; SIEGELOVÁ, J.; DUŠEK, J.; SOSÍKOVÁ, M.; HALBERG, F.

    2008-01-01

    In order to investigate any circannual and/or circaseptan variations in birth incidence and birth weight in Toda City (Japan), data on 4,411 consecutive births were obtained from the city’s Maternity Hospital between 1 Jan 1999 and 31 Dec 2001. Data were analysed by cosinor separately for babies with birth weights in given ranges, and separately for boys and girls born at different gestational ages. A circannual rhythm was detected with statistical significance (P=0.047) for birth incidence of all vaginal deliveries, with an acrophase in the fall. A similar result for caesarean sections was of borderline statistical significance. A circaseptan component with a relatively consistent acrophase around midweek was of borderline statistical significance for birth incidence in some of the groups investigated. About-yearly and about-weekly variations were also found to characterize birth weight in some of the groups investigated. PMID:18978949

  3. Development of classification model and QSAR model for predicting binding affinity of endocrine disrupting chemicals to human sex hormone-binding globulin.

    PubMed

    Liu, Huihui; Yang, Xianhai; Lu, Rui

    2016-08-01

    Disturbing the transport process is a crucial pathway for endocrine disrupting chemicals (EDCs) to disrupt endocrine function. However, this mechanism has not gotten enough attention, compared with that of hormone receptors and synthetase up to now, especially for the sex hormone transport process. In this study, we selected sex hormone-binding globulin (SHBG) and EDCs as a model system and the relative competing potency of a chemical with testosterone binding to SHBG (log RBA) as the endpoints, to develop classification models and quantitative structure-activity relationship (QSAR) models. With the classification model, a satisfactory model with nR09, nR10 and RDF155v as the most relevant variables was screened. Statistic results indicated that the model had the sensitivity, specificity, accuracy of 86.4%, 80.0%, 84.4% and 85.7%, 87.5%, 86.2% for the training set and validation set, respectively, highlighting a high classification performance of the model. With the QSAR model, a satisfactory model with statistical parameters, specifically, an adjusted determination coefficient (Radj(2)) of 0.810, a root mean square error (RMSE) of 0.616, a leave-one-out cross-validation squared correlation coefficient (QLOO(2)) of 0.777, a bootstrap method (QBOOT(2)) of 0.756, an external validation coefficient (Qext(2)) of 0.544 and a RMSEext of 0.859, were obtained, which implied satisfactory goodness of fit, robustness and predictive ability. The applicability domain of the current model covers a large number of structurally diverse chemicals, especially a few classes of nonsteroidal compounds. PMID:27156209

  4. Affinity chromatography: a historical perspective.

    PubMed

    Hage, David S; Matsuda, Ryan

    2015-01-01

    Affinity chromatography is one of the most selective and versatile forms of liquid chromatography for the separation or analysis of chemicals in complex mixtures. This method makes use of a biologically related agent as the stationary phase, which provides an affinity column with the ability to bind selectively and reversibly to a given target in a sample. This review examines the early work in this method and various developments that have lead to the current status of this technique. The general principles of affinity chromatography are briefly described as part of this discussion. Past and recent efforts in the generation of new binding agents, supports, and immobilization methods for this method are considered. Various applications of affinity chromatography are also summarized, as well as the influence this field has played in the creation of other affinity-based separation or analysis methods. PMID:25749941

  5. Quantum groups, braiding matrices and coset models

    SciTech Connect

    Itoyama, H.

    1989-07-01

    We discuss a few results on quantum groups in the context of rational conformal field theory with underlying affine Lie algebras. A vertex-height correspondence - a well-known procedure in solvable lattice models - is introduced in the WZW theory. This leads to a new definition of chiral vertex operator in which the zero mode is given by the q-Clebsch Gordan coefficients. Braiding matrices of coset models are found to factorize into those of the WZW theories. We briefly discuss the construction of the generators of the universal enveloping algebra in Toda field theories. 21 refs., 2 figs.

  6. Affinity Chromatography in Nonionic Detergent Solutions

    NASA Astrophysics Data System (ADS)

    Robinson, Jack B.; Strottmann, James M.; Wick, Donald G.; Stellwagen, Earle

    1980-10-01

    Anionic dye affinity chromatography is commonly unproductive in the presence of nonionic detergents used to extract particulate proteins. Using lactate dehydrogenase as a model protein, Cibacron blue F3GA as a model dye, and Triton X-100 as a model detergent, we find that the dye is encapsulated in nonionic detergent micelles, rendering the dye incapable of ligation with the enzyme. However, the dye can be liberated from the micelles without altering the nonionic detergent concentration by addition of an anionic detergent, such as deoxycholate or sodium dodecyl sulfate, forming mixed anionic/nonionic micelles that displace the anionic dye. Encapsulation of the anionic detergents prevents their activity as protein denaturants. These observations have been successfully translated to the dye affinity chromatography of a detergent extract of brain particulate cyclic nucleotide phosphodiesterase.

  7. Mutations of the cluster algebra of type {A}_{1}^{(1)} and the periodic discrete Toda lattice

    NASA Astrophysics Data System (ADS)

    Nobe, Atsushi

    2016-07-01

    A direct connection between two sequences of points, one of which is generated by seed mutations of the cluster algebra of type {A}1(1) and the other by time evolutions of the periodic discrete Toda lattice, is explicitly given. In this construction, each of them is realized as an orbit of a QRT map, and specialization of the parameters in the maps and appropriate choices of the initial points relate them. The connection with the periodic discrete Toda lattice enables us a geometric interpretation of the seed mutations of the cluster algebra of type {A}1(1) as an addition of points on an elliptic curve.

  8. Exploring the Origin of Differential Binding Affinities of Human Tubulin Isotypes αβII, αβIII and αβIV for DAMA-Colchicine Using Homology Modelling, Molecular Docking and Molecular Dynamics Simulations

    PubMed Central

    Panda, Dulal; Kunwar, Ambarish

    2016-01-01

    Tubulin isotypes are found to play an important role in regulating microtubule dynamics. The isotype composition is also thought to contribute in the development of drug resistance as tubulin isotypes show differential binding affinities for various anti-cancer agents. Tubulin isotypes αβII, αβIII and αβIV show differential binding affinity for colchicine. However, the origin of differential binding affinity is not well understood at the molecular level. Here, we investigate the origin of differential binding affinity of a colchicine analogue N-deacetyl-N-(2-mercaptoacetyl)-colchicine (DAMA-colchicine) for human αβII, αβIII and αβIV isotypes, employing sequence analysis, homology modeling, molecular docking, molecular dynamics simulation and MM-GBSA binding free energy calculations. The sequence analysis study shows that the residue compositions are different in the colchicine binding pocket of αβII and αβIII, whereas no such difference is present in αβIV tubulin isotypes. Further, the molecular docking and molecular dynamics simulations results show that residue differences present at the colchicine binding pocket weaken the bonding interactions and the correct binding of DAMA-colchicine at the interface of αβII and αβIII tubulin isotypes. Post molecular dynamics simulation analysis suggests that these residue variations affect the structure and dynamics of αβII and αβIII tubulin isotypes, which in turn affect the binding of DAMA-colchicine. Further, the binding free-energy calculation shows that αβIV tubulin isotype has the highest binding free-energy and αβIII has the lowest binding free-energy for DAMA-colchicine. The order of binding free-energy for DAMA-colchicine is αβIV ≃ αβII >> αβIII. Thus, our computational approaches provide an insight into the effect of residue variations on differential binding of αβII, αβIII and αβIV tubulin isotypes with DAMA-colchicine and may help to design new analogues with higher

  9. Exploring the Origin of Differential Binding Affinities of Human Tubulin Isotypes αβII, αβIII and αβIV for DAMA-Colchicine Using Homology Modelling, Molecular Docking and Molecular Dynamics Simulations.

    PubMed

    Kumbhar, Bajarang Vasant; Borogaon, Anubhaw; Panda, Dulal; Kunwar, Ambarish

    2016-01-01

    Tubulin isotypes are found to play an important role in regulating microtubule dynamics. The isotype composition is also thought to contribute in the development of drug resistance as tubulin isotypes show differential binding affinities for various anti-cancer agents. Tubulin isotypes αβII, αβIII and αβIV show differential binding affinity for colchicine. However, the origin of differential binding affinity is not well understood at the molecular level. Here, we investigate the origin of differential binding affinity of a colchicine analogue N-deacetyl-N-(2-mercaptoacetyl)-colchicine (DAMA-colchicine) for human αβII, αβIII and αβIV isotypes, employing sequence analysis, homology modeling, molecular docking, molecular dynamics simulation and MM-GBSA binding free energy calculations. The sequence analysis study shows that the residue compositions are different in the colchicine binding pocket of αβII and αβIII, whereas no such difference is present in αβIV tubulin isotypes. Further, the molecular docking and molecular dynamics simulations results show that residue differences present at the colchicine binding pocket weaken the bonding interactions and the correct binding of DAMA-colchicine at the interface of αβII and αβIII tubulin isotypes. Post molecular dynamics simulation analysis suggests that these residue variations affect the structure and dynamics of αβII and αβIII tubulin isotypes, which in turn affect the binding of DAMA-colchicine. Further, the binding free-energy calculation shows that αβIV tubulin isotype has the highest binding free-energy and αβIII has the lowest binding free-energy for DAMA-colchicine. The order of binding free-energy for DAMA-colchicine is αβIV ≃ αβII > αβIII. Thus, our computational approaches provide an insight into the effect of residue variations on differential binding of αβII, αβIII and αβIV tubulin isotypes with DAMA-colchicine and may help to design new analogues with higher

  10. Streptomycin affinity depends on 13 amino acids forming a loop in homology modelled ribosomal S12 protein (rpsL gene) of Lysinibacillus sphaericus DSLS5 associated with marine sponge (Tedania anhelans).

    PubMed

    Suriyanarayanan, Balasubramanian; Lakshmi, Praveena Pothuraju; Santhosh, Ramachandran Sarojini; Dhevendaran, Kandasamy; Priya, Balakrishnan; Krishna, Shivaani

    2016-06-01

    Streptomycin, an antibiotic used against microbial infections, inhibits the protein synthesis by binding to ribosomal protein S12, encoded by rpsL12 gene, and associated mutations cause streptomycin resistance. A streptomycin resistant, Lysinibacillus sphaericus DSLS5 (MIC >300 µg/mL for streptomycin), was isolated from a marine sponge (Tedania anhelans). The characterisation of rpsL12 gene showed a region having similarity to long terminal repeat sequences of murine lukemia virus which added 13 amino acids for loop formation in RpsL12; in addition, a K56R mutation which corresponds to K43R mutation present in streptomycin-resistant Escherichia coli is also present. The RpsL12 protein was modelled and compared with that of Lysinibacillus boronitolerans, Escherichia coli and Mycobacterium tuberculosis. The modelled proteins docked with streptomycin indicate compound had less affinity. The effect of loop on streptomycin resistance was analysed by constructing three different models of RpsL12 by, (i) removing both loop and mutation, (ii) removing the loop alone while retaining the mutation and (iii) without mutation having loop. The results showed that the presence of loop causes streptomycin resistance (decreases the affinity), and it further enhanced in the presence of mutation at 56th codon. Further study will help in understanding the evolution of streptomycin resistance in organisms. PMID:26198082

  11. DNA-binding affinity and anticancer activity of β-carboline-chalcone conjugates as potential DNA intercalators: Molecular modelling and synthesis.

    PubMed

    Shankaraiah, Nagula; Siraj, K P; Nekkanti, Shalini; Srinivasulu, Vunnam; Sharma, Pankaj; Senwar, Kishna Ram; Sathish, Manda; Vishnuvardhan, M V P S; Ramakrishna, Sistla; Jadala, Chetna; Nagesh, Narayana; Kamal, Ahmed

    2015-04-01

    A new series of DNA-interactive β-carboline-chalcone conjugates have been synthesized and evaluated for their in vitro cytotoxicity and DNA-binding affinity. It has been observed that most of these new hybrids have shown potent cytotoxic activities on A-549 (lung adenocarcinoma) cell lines with IC50 values lower than 10 μM. The hybrid 7b is more effective against some of the selected cancer cell lines with IC50 values less than 50 μM. In addition, compounds 7e, 7k, 7p-u has displayed significant elevation in ΔTm of DNA in comparison to Adriamycin, suggesting significant interaction and remarkable DNA stabilization. The DNA intercalation of these new hybrids has been investigated by fluorescence titration, DNA viscosity measurements, molecular docking as well as molecular dynamics and the results are in agreement with the thermal denaturation studies. PMID:25771335

  12. Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high-affinity receptor FcεRI.

    PubMed

    Sabban, Sari; Ye, Hongtu; Helm, Birgit

    2014-01-01

    The interaction of IgE with its high-affinity Fc receptor (FcεRI) followed by an antigenic challenge is the principal pathway in IgE mediated allergic reactions. As a consequence of the high affinity binding between IgE and FcεRI, along with the continuous production of IgE by B cells, allergies usually persist throughout life, with currently no permanent cure available. Horses, especially race horses, which are commonly inbred, are a species of mammals that are very prone to the development of hypersensitivity responses, which can seriously affect their performance. Physiological responses to allergic sensitization in horses mirror that observed in humans and dogs. In this paper we describe the development of an in situ assay system for the quantitative assessment of the release of mediators of the allergic response pertaining to the equine system. To this end, the gene encoding equine FcεRIα was transfected into and expressed onto the surface of parental Rat Basophil Leukemia (RBL-2H3.1) cells. The gene product of the transfected equine α-chain formed a functional receptor complex with the endogenous rat β- and γ-chains. The resultant assay system facilitated an assessment of the quantity of mediator secreted from equine FcεRIα transfected RBL-2H3.1 cells following sensitization with equine IgE and antigenic challenge using β-hexosaminidase release as a readout. Mediator release peaked at 36.68% ± 4.88% at 100 ng ml(-1) of antigen. This assay was modified from previous assays used to study human and canine allergic responses. We have also shown that this type of assay system has multiple applications for the development of diagnostic tools and the safety assessment of potential therapeutic intervention strategies in allergic disease. PMID:25406512

  13. Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high-affinity receptor FcɛRI.

    PubMed

    Sabban, Sari; Ye, Hongtu; Helm, Birgit

    2013-05-15

    The binding of immunoglobulin E (IgE) to its high-affinity receptor (FcɛRI) is the central protein interaction in IgE-mediated allergic reactions. The cross-linking of the IgE/FcɛRI complex, through cognate allergens, on the surface of mast cells and basophil cells results in mediator release, and thus leads to the symptoms of type I hypersensitivity responses in mammals. To develop a baseline value for subsequent equine anti-allergy drug and vaccine research, the interaction of equine IgE with its high-affinity FcɛRI receptor was investigated following the cloning and expression of equine IgE with specificity for NIP-HSA (4-hydroxy-5-iodo-3-nitrophenylacetic acid conjugated to human serum albumin). Receptor recognition and effector functions were assessed in Rat Basophil Leukemia (RBL-2H3.1) cells transfected with the α chain of equine and canine FcɛRI. Results obtained showed that the equine FcɛRI receptor recognizes both equine and canine IgE and supports similar β-hexosaminidase release levels from RBL cells transfected with equine FcɛRI, peaking at 36.68% at 100ngml(-1) antigen and 32.00% at 100ngml(-1) antigen respectively. Furthermore, the binding kinetics of the equine IgE to the equine FcɛRI receptor and the canine IgE to the same receptor was measured to be KA=6.33×10(9)M(-1) and KA=1.84×10(9)M(-1) respectively. This research established basic reagents and vitro assay systems to underpin the development of rational therapeutic intervention strategies to combat equine allergic manifestations. PMID:23485176

  14. Smooth big bounce from affine quantization

    NASA Astrophysics Data System (ADS)

    Bergeron, Hervé; Dapor, Andrea; Gazeau, Jean Pierre; Małkiewicz, Przemysław

    2014-04-01

    We examine the possibility of dealing with gravitational singularities on a quantum level through the use of coherent state or wavelet quantization instead of canonical quantization. We consider the Robertson-Walker metric coupled to a perfect fluid. It is the simplest model of a gravitational collapse, and the results obtained here may serve as a useful starting point for more complex investigations in the future. We follow a quantization procedure based on affine coherent states or wavelets built from the unitary irreducible representation of the affine group of the real line with positive dilation. The main issue of our approach is the appearance of a quantum centrifugal potential allowing for regularization of the singularity, essential self-adjointness of the Hamiltonian, and unambiguous quantum dynamical evolution.

  15. Potential Modes of Interaction of 9-Aminomethyl-9,10-dihydroanthracene (AMDA) Derivatives with the 5-HT2A Receptor: A Ligand Structure-Affinity Relationship, Receptor Mutagenesis and Receptor Modeling Investigation⊕

    PubMed Central

    Runyon, Scott P.; Mosier, Philip D.; Roth, Bryan L.; Glennon, Richard A.; Westkaemper, Richard B.

    2011-01-01

    The effects of 3-position substitution of 9-aminomethyl-9,10-dihydroanthracene (AMDA) on 5-HT2A receptor affinity were determined and compared to a parallel series of DOB-like 1-(2,5-dimethoxyphenyl)-2-aminopropanes substituted at the 4-position. The results were interpreted within the context of 5-HT2A receptor models that suggest that members of the DOB-like series can bind to the receptor in two distinct modes that correlate with the compounds’ functional activity. Automated ligand docking and molecular dynamics suggest that all of the AMDA derivatives, the parent of which is a 5-HT2A antagonist, bind in a fashion analogous to that for the sterically demanding antagonist DOB-like compounds. The failure of the F3406.52L mutation to adversely affect the affinity of AMDA and the 3-bromo derivative is consistent with the proposed modes of orientation. Evaluation of ligand-receptor complex models suggest that a valine/threonine exchange between the 5-HT2A and D2 receptors may be the origin of selectivity for AMDA and two substituted derivatives. PMID:18847250

  16. Overview of affinity tags for protein purification.

    PubMed

    Kimple, Michelle E; Sondek, John

    2004-09-01

    Addition of an affinity tag is a useful method for differentiating recombinant proteins expressed in bacterial and eukaryotic expression systems from the background of total cellular proteins, and for detecting protein-protein interactions. This overview describes the historical basis for the development of affinity tags, affinity tags that are commonly used today, how to choose an appropriate affinity tag for a particular purpose, and several recently developed affinity tag technologies that may prove useful in the near future. PMID:18429272

  17. Overview of affinity tags for protein purification.

    PubMed

    Kimple, Michelle E; Brill, Allison L; Pasker, Renee L

    2013-01-01

    Addition of an affinity tag is a useful method for differentiating recombinant proteins expressed in bacterial and eukaryotic expression systems from the background of total cellular proteins, as well as for detecting protein-protein interactions. This overview describes the historical basis for the development of affinity tags, affinity tags that are commonly used today, how to choose an appropriate affinity tag for a particular purpose, and several recently developed affinity tag technologies that may prove useful in the near future. PMID:24510596

  18. Affine Contractions on the Plane

    ERIC Educational Resources Information Center

    Celik, D.; Ozdemir, Y.; Ureyen, M.

    2007-01-01

    Contractions play a considerable role in the theory of fractals. However, it is not easy to find contractions which are not similitudes. In this study, it is shown by counter examples that an affine transformation of the plane carrying a given triangle onto another triangle may not be a contraction even if it contracts edges, heights or medians.…

  19. Quantifying Affinity among Chinese Dialects.

    ERIC Educational Resources Information Center

    Cheng, Chin-Chuan

    A study of the relationships between Chinese dialects based on a quantitative measure of dialect affinity is summarized. First, tone values in all the dialect localities available in the early 1970s were used to calculate the dialectal differences in terms of tone height with respect to the "yin and yang" split. In the late 1970s, calculations of…

  20. Theoretical proton affinity and fluoride affinity of nerve agent VX.

    PubMed

    Bera, Narayan C; Maeda, Satoshi; Morokuma, Keiji; Viggiano, Al A

    2010-12-23

    Proton affinity and fluoride affinity of nerve agent VX at all of its possible sites were calculated at the RI-MP2/cc-pVTZ//B3LYP/6-31G* and RI-MP2/aug-cc-pVTZ//B3LYP/6-31+G* levels, respectively. The protonation leads to various unique structures, with H(+) attached to oxygen, nitrogen, and sulfur atoms; among which the nitrogen site possesses the highest proton affinity of -ΔE ∼ 251 kcal/mol, suggesting that this is likely to be the major product. In addition some H(2), CH(4) dissociation as well as destruction channels have been found, among which the CH(4) + [Et-O-P(═O)(Me)-S-(CH(2))(2)-N(+)(iPr)═CHMe] product and the destruction product forming Et-O-P(═O)(Me)-SMe + CH(2)═N(+)(iPr)(2) are only 9 kcal/mol less stable than the most stable N-protonated product. For fluoridization, the S-P destruction channel to give Et-O-P(═O)(Me)(F) + [S-(CH(2))(2)-N-(iPr)(2)](-) is energetically the most favorable, with a fluoride affinity of -ΔE ∼ 44 kcal. Various F(-) ion-molecule complexes are also found, with the one having F(-) interacting with two hydrogen atoms in different alkyl groups to be only 9 kcal/mol higher than the above destruction product. These results suggest VX behaves quite differently from surrogate systems. PMID:21117653

  1. Bidirectional Elastic Image Registration Using B-Spline Affine Transformation

    PubMed Central

    Gu, Suicheng; Meng, Xin; Sciurba, Frank C.; Wang, Chen; Kaminski, Naftali; Pu, Jiantao

    2014-01-01

    A registration scheme termed as B-spline affine transformation (BSAT) is presented in this study to elastically align two images. We define an affine transformation instead of the traditional translation at each control point. Mathematically, BSAT is a generalized form of the affine transformation and the traditional B-Spline transformation (BST). In order to improve the performance of the iterative closest point (ICP) method in registering two homologous shapes but with large deformation, a bi-directional instead of the traditional unidirectional objective / cost function is proposed. In implementation, the objective function is formulated as a sparse linear equation problem, and a sub-division strategy is used to achieve a reasonable efficiency in registration. The performance of the developed scheme was assessed using both two-dimensional (2D) synthesized dataset and three-dimensional (3D) volumetric computed tomography (CT) data. Our experiments showed that the proposed B-spline affine model could obtain reasonable registration accuracy. PMID:24530210

  2. Bidirectional elastic image registration using B-spline affine transformation.

    PubMed

    Gu, Suicheng; Meng, Xin; Sciurba, Frank C; Ma, Hongxia; Leader, Joseph; Kaminski, Naftali; Gur, David; Pu, Jiantao

    2014-06-01

    A registration scheme termed as B-spline affine transformation (BSAT) is presented in this study to elastically align two images. We define an affine transformation instead of the traditional translation at each control point. Mathematically, BSAT is a generalized form of the affine transformation and the traditional B-spline transformation (BST). In order to improve the performance of the iterative closest point (ICP) method in registering two homologous shapes but with large deformation, a bidirectional instead of the traditional unidirectional objective/cost function is proposed. In implementation, the objective function is formulated as a sparse linear equation problem, and a sub-division strategy is used to achieve a reasonable efficiency in registration. The performance of the developed scheme was assessed using both two-dimensional (2D) synthesized dataset and three-dimensional (3D) volumetric computed tomography (CT) data. Our experiments showed that the proposed B-spline affine model could obtain reasonable registration accuracy. PMID:24530210

  3. Fatigue damage prognosis using affine arithmetic

    NASA Astrophysics Data System (ADS)

    Gbaguidi, Audrey; Kim, Daewon

    2014-02-01

    Among the essential steps to be taken in structural health monitoring systems, damage prognosis would be the field that is least investigated due to the complexity of the uncertainties. This paper presents the possibility of using Affine Arithmetic for uncertainty propagation of crack damage in damage prognosis. The structures examined are thin rectangular plates made of titanium alloys with central mode I cracks and a composite plate with an internal delamination caused by mixed mode I and II fracture modes, under a harmonic uniaxial loading condition. The model-based method for crack growth rates are considered using the Paris Erdogan law model for the isotropic plates and the delamination growth law model proposed by Kardomateas for the composite plate. The parameters for both models are randomly taken and their uncertainties are considered as defined by an interval instead of a probability distribution. A Monte Carlo method is also applied to check whether Affine Arithmetic (AA) leads to tight bounds on the lifetime of the structure.

  4. Lectin affinity chromatography of glycolipids

    SciTech Connect

    Torres, B.V.; Smith, D.F.

    1987-05-01

    Since glycolipids (GLs) are either insoluble or form mixed micelles in water, lectin affinity chromatography in aqueous systems has not been applied to their separation. They have overcome this problem by using tetrahydrofuran (THF) in the mobile phase during chromatography. Affinity columns prepared with the GalNAc-specific Helix pomatia agglutinin (HPA) and equilibrated in THF specifically bind the (/sup 3/H)oligosaccharide derived from Forssman GL indicating that the immobilized HPA retained its carbohydrate-binding specificity in this solvent. Intact Forssman GL was bound by the HPA-column equilibrated in THF and was specifically eluted with 0.1 mg/ml GalNAc in THF. Purification of the Forssman GL was achieved when a crude lipid extract of sheep erythrocyte membranes was applied to the HPA-column in THF. Non-specifically bound GLs were eluted from the column using a step gradient of aqueous buffer in THF, while the addition of GalNAc was required to elute the specifically bound GLs. Using this procedure the A-active GLs were purified from a crude lipid extract of type A human erythrocytes in a single chromatographic step. The use of solvents that maintain carbohydrate-binding specificity and lipid solubility will permit the application of affinity chromatography on immobilized carbohydrate-binding proteins to intact GLs.

  5. Experimental and theoretical binding affinity between polyvinylpolypyrrolidone and selected phenolic compounds from food matrices.

    PubMed

    Durán-Lara, Esteban F; López-Cortés, Xaviera A; Castro, Ricardo I; Avila-Salas, Fabián; González-Nilo, Fernando D; Laurie, V Felipe; Santos, Leonardo S

    2015-02-01

    Polyvinylpolypyrrolidone (PVPP) is a fining agent, widely used in winemaking and brewing, whose mode of action in removing phenolic compounds has not been fully characterised. The aim of this study was to evaluate the experimental and theoretical binding affinity of PVPP towards six phenolic compounds representing different types of phenolic species. The interaction between PVPP and phenolics was evaluated in model solutions, where hydroxyl groups, hydrophobic bonding and steric hindrance were characterised. The results of the study indicated that PVPP exhibits high affinity for quercetin and catechin, moderate affinity for epicatechin, gallic acid and lower affinity for 4-methylcatechol and caffeic acid. The affinity has a direct correlation with the hydroxylation degree of each compound. The results show that the affinity of PVPP towards phenols is related with frontier orbitals. This work demonstrates a direct correlation between the experimental affinity and the interaction energy calculations obtained through computational chemistry methods. PMID:25172736

  6. ESTIMATION OF ELECTRON AFFINITY BASED ON STRUCTURE ACTIVITY RELATIONSHIPS

    EPA Science Inventory

    Electron affinity for a wide range of organic molecules was calculated from molecular structure using the chemical reactivity models developed in SPARC. hese models are based on fundamental chemical structure theory applied to the prediction of chemical reactivities for organic m...

  7. Immobilized metal ion affinity chromatography.

    PubMed

    Yip, T T; Hutchens, T W

    1992-01-01

    Immobilized metal ion affinity chromatography (IMAC) (1,2) is also referred to as metal chelate chromatography, metal ion interaction chromatography, and ligand-exchange chromatography. We view this affinity separation technique as an intermediate between highly specific, high-affinity bioaffinity separation methods, and wider spectrum, low-specificity adsorption methods, such as ion exchange. The IMAC stationary phases are designed to chelate certain metal ions that have selectivity for specific groups (e.g., His residues) in peptides (e.g., 3-7) and on protein surfaces (8-13). The number of stationary phases that can be synthesized for efficient chelation of metal ions is unlimited, but the critical consideration is that there must be enough exposure of the metal ion to interact with the proteins, preferably in a biospecific manner. Several examples are presented in Fig. 1. The challenge to produce new immobilized chelating groups, including protein surface metal-binding domains (14,15) is being explored continuously. Table 1 presents a list of published procedures for the synthesis and use of stationary phases with immobilized chelating groups. This is by no means exhaustive, and is intended only to give an idea of the scope and versatility of IMAC. Fig. 1 Schematic illustration of several types of immobilized metal-chelating groups, including, iminodiacetate (IDA), tris(carboxymethyl) ethylenediamine (TED), and the metal-binding peptides (GHHPH)(n)G (where n = 1,2,3, and 5) (14,15). Table 1 Immobilized Chelating Groups and Metal Ions Used for Immobilized Metal Ion Affinity Chromatography Chelating group Suitable metal ions Reference Commercial source Immodiacetate Transitional1,2 Pharmacia LKB Pierce Sigma Boehringer Mannheim TosoHaas 2-Hydroxy-3[N-(2- pyrtdylmethyl) glycme]propyl Transitional3 Not available ?-Alky1 mtrilo triacetic acid Transitional4 Not available Carboxymethylated asparhc acid Ca(II)13 Not available Tris (carboxy- methyl) ethylene Diamme

  8. Indian craniometric variability and affinities.

    PubMed

    Raghavan, Pathmanathan; Bulbeck, David; Pathmanathan, Gayathiri; Rathee, Suresh Kanta

    2013-01-01

    Recently published craniometric and genetic studies indicate a predominantly indigenous ancestry of Indian populations. We address this issue with a fuller coverage of Indian craniometrics than any done before. We analyse metrical variability within Indian series, Indians' sexual dimorphism, differences between northern and southern Indians, index-based differences of Indian males from other series, and Indians' multivariate affinities. The relationship between a variable's magnitude and its variability is log-linear. This relationship is strengthened by excluding cranial fractions and series with a sample size less than 30. Male crania are typically larger than female crania, but there are also shape differences. Northern Indians differ from southern Indians in various features including narrower orbits and less pronounced medial protrusion of the orbits. Indians resemble Veddas in having small crania and similar cranial shape. Indians' wider geographic affinities lie with "Caucasoid" populations to the northwest, particularly affecting northern Indians. The latter finding is confirmed from shape-based Mahalanobis-D distances calculated for the best sampled male and female series. Demonstration of a distinctive South Asian craniometric profile and the intermediate status of northern Indians between southern Indians and populations northwest of India confirm the predominantly indigenous ancestry of northern and especially southern Indians. PMID:24455409

  9. Indian Craniometric Variability and Affinities

    PubMed Central

    Raghavan, Pathmanathan; Bulbeck, David; Pathmanathan, Gayathiri; Rathee, Suresh Kanta

    2013-01-01

    Recently published craniometric and genetic studies indicate a predominantly indigenous ancestry of Indian populations. We address this issue with a fuller coverage of Indian craniometrics than any done before. We analyse metrical variability within Indian series, Indians' sexual dimorphism, differences between northern and southern Indians, index-based differences of Indian males from other series, and Indians' multivariate affinities. The relationship between a variable's magnitude and its variability is log-linear. This relationship is strengthened by excluding cranial fractions and series with a sample size less than 30. Male crania are typically larger than female crania, but there are also shape differences. Northern Indians differ from southern Indians in various features including narrower orbits and less pronounced medial protrusion of the orbits. Indians resemble Veddas in having small crania and similar cranial shape. Indians' wider geographic affinities lie with “Caucasoid” populations to the northwest, particularly affecting northern Indians. The latter finding is confirmed from shape-based Mahalanobis-D distances calculated for the best sampled male and female series. Demonstration of a distinctive South Asian craniometric profile and the intermediate status of northern Indians between southern Indians and populations northwest of India confirm the predominantly indigenous ancestry of northern and especially southern Indians. PMID:24455409

  10. Low-Affinity Memory CD8+ T Cells Mediate Robust Heterologous Immunity.

    PubMed

    Krummey, Scott M; Martinez, Ryan J; Andargachew, Rakieb; Liu, Danya; Wagener, Maylene; Kohlmeier, Jacob E; Evavold, Brian D; Larsen, Christian P; Ford, Mandy L

    2016-03-15

    Heterologous immunity is recognized as a significant barrier to transplant tolerance. Whereas it has been established that pathogen-elicited memory T cells can have high or low affinity for cross-reactive allogeneic peptide-MHC, the role of TCR affinity during heterologous immunity has not been explored. We established a model with which to investigate the impact of TCR-priming affinity on memory T cell populations following a graft rechallenge. In contrast to high-affinity priming, low-affinity priming elicited fully differentiated memory T cells with a CD45RB(hi) status. High CD45RB status enabled robust secondary responses in vivo, as demonstrated by faster graft rejection kinetics and greater proliferative responses. CD45RB blockade prolonged graft survival in low affinity-primed mice, but not in high affinity-primed mice. Mechanistically, low affinity-primed memory CD8(+) T cells produced more IL-2 and significantly upregulated IL-2Rα expression during rechallenge. We found that CD45RB(hi) status was also a stable marker of priming affinity within polyclonal CD8(+) T cell populations. Following high-affinity rechallenge, low affinity-primed CD45RB(hi) cells became CD45RB(lo), demonstrating that CD45RB status acts as an affinity-based differentiation switch on CD8(+) T cells. Thus, these data establish a novel mechanism by which CD45 isoforms tune low affinity-primed memory CD8(+) T cells to become potent secondary effectors following heterologous rechallenge. These findings have direct implications for allogeneic heterologous immunity by demonstrating that despite a lower precursor frequency, low-affinity priming is sufficient to generate memory cells that mediate potent secondary responses against a cross-reactive graft challenge. PMID:26864034

  11. Background correction using dinucleotide affinities improves the performance of GCRMA

    PubMed Central

    Gharaibeh, Raad Z; Fodor, Anthony A; Gibas, Cynthia J

    2008-01-01

    Background High-density short oligonucleotide microarrays are a primary research tool for assessing global gene expression. Background noise on microarrays comprises a significant portion of the measured raw data, which can have serious implications for the interpretation of the generated data if not estimated correctly. Results We introduce an approach to calculate probe affinity based on sequence composition, incorporating nearest-neighbor (NN) information. Our model uses position-specific dinucleotide information, instead of the original single nucleotide approach, and adds up to 10% to the total variance explained (R2) when compared to the previously published model. We demonstrate that correcting for background noise using this approach enhances the performance of the GCRMA preprocessing algorithm when applied to control datasets, especially for detecting low intensity targets. Conclusion Modifying the previously published position-dependent affinity model to incorporate dinucleotide information significantly improves the performance of the model. The dinucleotide affinity model enhances the detection of differentially expressed genes when implemented as a background correction procedure in GeneChip preprocessing algorithms. This is conceptually consistent with physical models of binding affinity, which depend on the nearest-neighbor stacking interactions in addition to base-pairing. PMID:18947404

  12. Increased hemoglobin O2 affinity protects during acute hypoxia.

    PubMed

    Yalcin, Ozlem; Cabrales, Pedro

    2012-08-01

    Acclimatization to hypoxia requires time to complete the adaptation mechanisms that influence oxygen (O(2)) transport and O(2) utilization. Although decreasing hemoglobin (Hb) O(2) affinity would favor the release of O(2) to the tissues, increasing Hb O(2) affinity would augment arterial O(2) saturation during hypoxia. This study was designed to test the hypothesis that pharmacologically increasing the Hb O(2) affinity will augment O(2) transport during severe hypoxia (10 and 5% inspired O(2)) compared with normal Hb O(2) affinity. RBC Hb O(2) affinity was increased by infusion of 20 mg/kg of 5-hydroxymethyl-2-furfural (5HMF). Control animals received only the vehicle. The effects of increasing Hb O(2) affinity were studied in the hamster window chamber model, in terms of systemic and microvascular hemodynamics and partial pressures of O(2) (Po(2)). Pimonidazole binding to hypoxic areas of mice heart and brain was also studied. 5HMF decreased the Po(2) at which the Hb is 50% saturated with O(2) by 12.6 mmHg. During 10 and 5% O(2) hypoxia, 5HMF increased arterial blood O(2) saturation by 35 and 48% from the vehicle group, respectively. During 5% O(2) hypoxia, blood pressure and heart rate were 58 and 30% higher for 5HMF compared with the vehicle. In addition, 5HMF preserved microvascular blood flow, whereas blood flow decreased to 40% of baseline in the vehicle group. Consequently, perivascular Po(2) was three times higher in the 5HMF group compared with the control group at 5% O(2) hypoxia. 5HMF also reduced heart and brain hypoxic areas in mice. Therefore, increased Hb O(2) affinity resulted in hemodynamics and oxygenation benefits during severe hypoxia. This acute acclimatization process may have implications in survival during severe environmental hypoxia when logistic constraints prevent chronic acclimatization. PMID:22636677

  13. Increased hemoglobin O2 affinity protects during acute hypoxia

    PubMed Central

    Yalcin, Ozlem

    2012-01-01

    Acclimatization to hypoxia requires time to complete the adaptation mechanisms that influence oxygen (O2) transport and O2 utilization. Although decreasing hemoglobin (Hb) O2 affinity would favor the release of O2 to the tissues, increasing Hb O2 affinity would augment arterial O2 saturation during hypoxia. This study was designed to test the hypothesis that pharmacologically increasing the Hb O2 affinity will augment O2 transport during severe hypoxia (10 and 5% inspired O2) compared with normal Hb O2 affinity. RBC Hb O2 affinity was increased by infusion of 20 mg/kg of 5-hydroxymethyl-2-furfural (5HMF). Control animals received only the vehicle. The effects of increasing Hb O2 affinity were studied in the hamster window chamber model, in terms of systemic and microvascular hemodynamics and partial pressures of O2 (Po2). Pimonidazole binding to hypoxic areas of mice heart and brain was also studied. 5HMF decreased the Po2 at which the Hb is 50% saturated with O2 by 12.6 mmHg. During 10 and 5% O2 hypoxia, 5HMF increased arterial blood O2 saturation by 35 and 48% from the vehicle group, respectively. During 5% O2 hypoxia, blood pressure and heart rate were 58 and 30% higher for 5HMF compared with the vehicle. In addition, 5HMF preserved microvascular blood flow, whereas blood flow decreased to 40% of baseline in the vehicle group. Consequently, perivascular Po2 was three times higher in the 5HMF group compared with the control group at 5% O2 hypoxia. 5HMF also reduced heart and brain hypoxic areas in mice. Therefore, increased Hb O2 affinity resulted in hemodynamics and oxygenation benefits during severe hypoxia. This acute acclimatization process may have implications in survival during severe environmental hypoxia when logistic constraints prevent chronic acclimatization. PMID:22636677

  14. Affine hypersurfaces with parallel difference tensor relative to affine α-connection

    NASA Astrophysics Data System (ADS)

    Li, Cece

    2014-12-01

    Li and Zhang (2014) studied affine hypersurfaces of R n + 1 with parallel difference tensor relative to the affine α-connection ∇ (α), and characterized the generalized Cayley hypersurfaces by K n - 1 ≠ 0 and ∇ (α) K = 0 for some nonzero constant α, where the affine α-connection ∇ (α) of information geometry was introduced on affine hypersurface. In this paper, by a slightly different method we continue to study affine hypersurfaces with ∇ (α) K = 0, if α = 0 we further assume that the Pick invariant vanishes and affine metric is of constant sectional curvature. It is proved that they are either hyperquadrics or improper affine hypersphere with flat indefinite affine metric, the latter can be locally given as a graph of a polynomial of at most degree n + 1 with constant Hessian determinant. In particular, if the affine metric is definite, Lorentzian, or its negative index is 2, we complete the classification of such hypersurfaces.

  15. GAB(A) receptors present higher affinity and modified subunit composition in spinal motor neurons from a genetic model of amyotrophic lateral sclerosis.

    PubMed

    Carunchio, Irene; Mollinari, Cristiana; Pieri, Massimo; Merlo, Daniela; Zona, Cristina

    2008-10-01

    Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by the selective degeneration of motor neurons in the spinal cord, brainstem and cerebral cortex. In this study we have analysed the electrophysiological properties of GABA(A) receptors and GABA(A) alpha1 and alpha2 subunits expression in spinal motor neurons in culture obtained from a genetic model of ALS (G93A) and compared with transgenic wild type SOD1 (SOD1) and their corresponding non transgenic litter mates (Control). Although excitotoxic motor neuron death has been extensively studied in relation to Ca(2+)-dependent processes, strong evidence indicates that excitotoxic cell death is also remarkably dependent on Cl(-) ions and on GABA(A) receptor activation. In this study we have analysed the electrophysiological properties of GABA(A) receptors and the expression of GABA(A)alpha(1) and alpha(2) subunits in cultured motor neurons obtained from a genetic model of amyotrophic lateral sclerosis (G93A) and compared them with transgenic wild-type Cu,Zn superoxide dismutase and their corresponding non-transgenic littermates (Control). In all tested motor neurons, the application of gamma-aminobutyric acid (GABA) (0.5-100 mum) evoked an inward current that was reversibly blocked by bicuculline (100 mum), thus indicating that it was mediated by the activation of GABA(A) receptors. Our results indicate that the current density at high GABA concentrations is similar in control, Cu,Zn superoxide dismutase and G93A motor neurons. However, the dose-response curve significantly shifted toward lower concentration values in G93A motor neurons and the extent of desensitization also increased in these neurons. Finally, multiplex single-cell real-time polymerase chain reaction and immunofluorescence revealed that the amount of GABA(A)alpha(1) subunit was significantly increased in G93A motor neurons, whereas the levels of alpha(2) subunit were unchanged. These data show that the functionality and

  16. The maximal affinity of ligands

    PubMed Central

    Kuntz, I. D.; Chen, K.; Sharp, K. A.; Kollman, P. A.

    1999-01-01

    We explore the question of what are the best ligands for macromolecular targets. A survey of experimental data on a large number of the strongest-binding ligands indicates that the free energy of binding increases with the number of nonhydrogen atoms with an initial slope of ≈−1.5 kcal/mol (1 cal = 4.18 J) per atom. For ligands that contain more than 15 nonhydrogen atoms, the free energy of binding increases very little with relative molecular mass. This nonlinearity is largely ascribed to nonthermodynamic factors. An analysis of the dominant interactions suggests that van der Waals interactions and hydrophobic effects provide a reasonable basis for understanding binding affinities across the entire set of ligands. Interesting outliers that bind unusually strongly on a per atom basis include metal ions, covalently attached ligands, and a few well known complexes such as biotin–avidin. PMID:10468550

  17. The crystal structure of oxy hemoglobin from high oxygen affinity bird emu (Dromaius novaehollandiae).

    PubMed

    Mohamed Abubakkar, Mohamed H; Saraboji, Kadhirvel; Ponnuswamy, Mon Nanjappa G

    2014-01-01

    Hemoglobin is an honorary enzyme, a two-way respiratory carrier, transporting oxygen from the lungs to the tissues and facilitating the return transport of carbon dioxide. Hemoglobin has high affinity for oxygen and low affinity for carbon dioxide and other substances in the arterial circulation, whereas in the venous circulation these relative affinities are upturned. The oxygen affinity of hemoglobin increases with the fall in temperature and decreases with the increase in pH and 2, 3-bisphosphoglycerate; point mutations also affect the tetrameric arrangement and alter the oxygen affinity. Though several studies have revealed the specific reasons for the adaptation of increased oxygen affinity of avian hemoglobins at high-altitudes, further structural insights on hemoglobins from high oxygen affinity species are required to understand the detailed oxygen adaptation at the molecular level. Herein, we describe the structural investigation of hemoglobin from emu (Dromaius novaehollandiae), a high oxygen affinity bird. Hemoglobin from emu was purified using anion-exchange chromatography, crystallized and determined the structure in the oxy form at a resolution of 2.3 Å; the R-factor of the model was 19.2%. The structure was compared with other oxy hemoglobins of high oxygen affinity avian species; significant changes are noted at intra-subunit contacts which provide the clues for increased oxygen affinity of emu hemoglobin. PMID:25146185

  18. Affine generalization of the Komar complex of general relativity

    NASA Astrophysics Data System (ADS)

    Mielke, Eckehard W.

    2001-02-01

    On the basis of the ``on shell'' Noether identities of the metric-affine gauge approach of gravity, an affine superpotential is derived which comprises the energy- and angular-momentum content of exact solutions. In the special case of general relativity (GR) or its teleparallel equivalent, the Komar or Freud complex, respectively, are recovered. Applying this to the spontaneously broken anti-de Sitter gauge model of McDowell and Mansouri with an induced Euler term automatically yields the correct mass and spin of the Kerr-AdS solution of GR with a (induced) cosmological constant without the factor two discrepancy of the Komar formula.

  19. Statistical theory of chromatography: new outlooks for affinity chromatography.

    PubMed Central

    Denizot, F C; Delaage, M A

    1975-01-01

    We have developed further the statistical approach to chromatography initiated by Giddings and Eyring, and applied it to affinity chromatography. By means of a convenient expression of moments the convergence towards the Laplace-Gauss distribution has been established. The Gaussian character is not preserved if other causes of dispersion are taken into account, but expressions of moments can be obtained in a generalized form. A simple procedure is deduced for expressing the fundamental constants of the model in terms of purely experimental quantities. Thus, affinity chromatography can be used to determine rate constants of association and dissociation in a range considered as the domain of the stopped-flow methods. PMID:1061072

  20. Dynamic output feedback H ∞ control for affine fuzzy systems

    NASA Astrophysics Data System (ADS)

    Wang, Huimin; Yang, Guang-Hong

    2013-06-01

    This article investigates the problem of designing H ∞ dynamic output feedback controllers for nonlinear systems, which are described by affine fuzzy models. The system outputs have been chosen as premise variables, which can guarantee that the plant and the controller always switch to the same region. By using a piecewise Lyapunov function and adding slack matrix variables, a piecewise-affine dynamic output feedback controller design method is obtained in the formulation of linear matrix inequalities (LMIs), which can be efficiently solved numerically. In contrast to the existing work, the proposed approach needs less LMI constraints and leads to less conservatism. Finally, numerical examples illustrate the effectiveness of the new result.

  1. Wetting on rough self-affine surfaces

    NASA Astrophysics Data System (ADS)

    Palasantzas, George

    1995-05-01

    In this paper, we present a general investigation of the effective potential for complete wetting on self-affine rough surfaces. The roughness effect is investigated by means of the height-height correlation model in Fourier space ~(1+aξ2q2)-1-H. The parameters H and ξ are, respectively, the roughness exponent and the substrate in-plane correlation length. It is observed that the effect of H on the free interface profile is significant for ξ>ξ) regime is characterized by a power-law scaling ~Y-2.

  2. Protein Complex Purification by Affinity Capture.

    PubMed

    LaCava, John; Fernandez-Martinez, Javier; Hakhverdyan, Zhanna; Rout, Michael P

    2016-01-01

    Affinity capture has become a powerful technique for consistently purifying endogenous protein complexes, facilitating biochemical and biophysical assays on otherwise inaccessible biological assemblies, and enabling broader interactomic exploration. For this procedure, cells are broken and their contents separated and extracted into a solvent, permitting access to target macromolecular complexes thus released in solution. The complexes are specifically enriched from the extract onto a solid medium coupled with an affinity reagent-usually an antibody-that recognizes the target either directly or through an appended affinity tag, allowing subsequent characterization of the complex. Here, we discuss approaches and considerations for purifying endogenous yeast protein complexes by affinity capture. PMID:27371601

  3. A Novel Vertex Affinity for Community Detection

    SciTech Connect

    Yoo, Andy; Sanders, Geoffrey; Henson, Van; Vassilevski, Panayot

    2015-10-05

    We propose a novel vertex affinity measure in this paper. The new vertex affinity quantifies the proximity between two vertices in terms of their clustering strength and is ideal for such graph analytics applications as community detection. We also developed a framework that combines simple graph searches and resistance circuit formulas to compute the vertex affinity efficiently. We study the properties of the new affinity measure empirically in comparison to those of other popular vertex proximity metrics. Our results show that the existing metrics are ill-suited for community detection due to their lack of fundamental properties that are essential for correctly capturing inter- and intra-cluster vertex proximity.

  4. Structural determinants of sigma receptor affinity

    SciTech Connect

    Largent, B.L.; Wikstroem, H.G.; Gundlach, A.L.; Snyder, S.H.

    1987-12-01

    The structural determinants of sigma receptor affinity have been evaluated by examining a wide range of compounds related to opioids, neuroleptics, and phenylpiperidine dopaminergic structures for affinity at sigma receptor-binding sites labeled with (+)-(/sup 3/H)3-PPP. Among opioid compounds, requirements for sigma receptor affinity differ strikingly from the determinants of affinity for conventional opiate receptors. Sigma sites display reverse stereoselectivity to classical opiate receptors. Multi-ringed opiate-related compounds such as morphine and naloxone have negligible affinity for sigma sites, with the highest sigma receptor affinity apparent for benzomorphans which lack the C ring of opioids. Highest affinity among opioids and other compounds occurs with more lipophilic N-substituents. This feature is particularly striking among the 3-PPP derivatives as well as the opioids. The butyrophenone haloperidol is the most potent drug at sigma receptors we have detected. Among the series of butyrophenones, receptor affinity is primarily associated with the 4-phenylpiperidine moiety. Conformational calculations for various compounds indicate a fairly wide range of tolerance for distances between the aromatic ring and the amine nitrogen, which may account for the potency at sigma receptors of structures of considerable diversity. Among the wide range of structures that bind to sigma receptor-binding sites, the common pharmacophore associated with high receptor affinity is a phenylpiperidine with a lipophilic N-substituent.

  5. Compact noncontraction semigroups of affine operators

    NASA Astrophysics Data System (ADS)

    Voynov, A. S.; Protasov, V. Yu

    2015-07-01

    We analyze compact multiplicative semigroups of affine operators acting in a finite-dimensional space. The main result states that every such semigroup is either contracting, that is, contains elements of arbitrarily small operator norm, or all its operators share a common invariant affine subspace on which this semigroup is contracting. The proof uses functional difference equations with contraction of the argument. We look at applications to self-affine partitions of convex sets, the investigation of finite affine semigroups and the proof of a criterion of primitivity for nonnegative matrix families. Bibliography: 32 titles.

  6. Evolution based on chromosome affinity from a network perspective

    NASA Astrophysics Data System (ADS)

    Monteiro, R. L. S.; Fontoura, J. R. A.; Carneiro, T. K. G.; Moret, M. A.; Pereira, H. B. B.

    2014-06-01

    Recent studies have focused on models to simulate the complex phenomenon of evolution of species. Several studies have been performed with theoretical models based on Darwin's theories to associate them with the actual evolution of species. However, none of the existing models include the affinity between individuals using network properties. In this paper, we present a new model based on the concept of affinity. The model is used to simulate the evolution of species in an ecosystem composed of individuals and their relationships. We propose an evolutive algorithm that incorporates the degree centrality and efficiency network properties to perform the crossover process and to obtain the network topology objective, respectively. Using a real network as a starting point, we simulate its evolution and compare its results with the results of 5788 computer-generated networks.

  7. Structure of classical affine and classical affine fractional W-algebras

    SciTech Connect

    Suh, Uhi Rinn

    2015-01-15

    We introduce a classical BRST complex (See Definition 3.2.) and show that one can construct a classical affine W-algebra via the complex. This definition clarifies that classical affine W-algebras can be considered as quasi-classical limits of quantum affine W-algebras. We also give a definition of a classical affine fractional W-algebra as a Poisson vertex algebra. As in the classical affine case, a classical affine fractional W-algebra has two compatible λ-brackets and is isomorphic to an algebra of differential polynomials as a differential algebra. When a classical affine fractional W-algebra is associated to a minimal nilpotent, we describe explicit forms of free generators and compute λ-brackets between them. Provided some assumptions on a classical affine fractional W-algebra, we find an infinite sequence of integrable systems related to the algebra, using the generalized Drinfel’d and Sokolov reduction.

  8. Scaling analysis of affinity propagation.

    PubMed

    Furtlehner, Cyril; Sebag, Michèle; Zhang, Xiangliang

    2010-06-01

    We analyze and exploit some scaling properties of the affinity propagation (AP) clustering algorithm proposed by Frey and Dueck [Science 315, 972 (2007)]. Following a divide and conquer strategy we setup an exact renormalization-based approach to address the question of clustering consistency, in particular, how many cluster are present in a given data set. We first observe that the divide and conquer strategy, used on a large data set hierarchically reduces the complexity O(N2) to O(N((h+2)/(h+1))) , for a data set of size N and a depth h of the hierarchical strategy. For a data set embedded in a d -dimensional space, we show that this is obtained without notably damaging the precision except in dimension d=2 . In fact, for d larger than 2 the relative loss in precision scales such as N((2-d)/(h+1)d). Finally, under some conditions we observe that there is a value s* of the penalty coefficient, a free parameter used to fix the number of clusters, which separates a fragmentation phase (for ss*) of the underlying hidden cluster structure. At this precise point holds a self-similarity property which can be exploited by the hierarchical strategy to actually locate its position, as a result of an exact decimation procedure. From this observation, a strategy based on AP can be defined to find out how many clusters are present in a given data set. PMID:20866473

  9. Affinity-based thermoresponsive precipitation of proteins modified with polymer-binding peptides.

    PubMed

    Suzuki, Seigo; Sawada, Toshiki; Ishizone, Takashi; Serizawa, Takeshi

    2016-04-14

    A 12-mer peptide with an affinity for the meso diad sequence of poly(N-isopropylacrylamide) (PNIPAM) was identified through affinity-based peptide screening. A model protein (i.e., human serum albumin (HSA)) chemically modified with the peptide was successfully precipitated with PNIPAM above the lower critical solution temperature (LCST) of PNIPAM. PMID:26996430

  10. Methods for Improving Aptamer Binding Affinity.

    PubMed

    Hasegawa, Hijiri; Savory, Nasa; Abe, Koichi; Ikebukuro, Kazunori

    2016-01-01

    Aptamers are single stranded oligonucleotides that bind a wide range of biological targets. Although aptamers can be isolated from pools of random sequence oligonucleotides using affinity-based selection, aptamers with high affinities are not always obtained. Therefore, further refinement of aptamers is required to achieve desired binding affinities. The optimization of primary sequences and stabilization of aptamer conformations are the main approaches to refining the binding properties of aptamers. In particular, sequence optimization using combined in silico sequence recombinations and in vitro functional evaluations is effective for the improvement of binding affinities, however, the binding affinities of aptamers are limited by the low hydrophobicity of nucleic acids. Accordingly, introduction of hydrophobic moieties into aptamers expands the diversity of interactions between aptamers and targets. Moreover, construction of multivalent aptamers by connecting aptamers that recognize distinct epitopes is an attractive approach to substantial increases in binding affinity. In addition, binding affinities can be tuned by optimizing the scaffolds of multivalent constructs. In this review, we summarize the various techniques for improving the binding affinities of aptamers. PMID:27043498

  11. Affine root systems and dual numbers

    NASA Astrophysics Data System (ADS)

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

    The root systems in Carroll spaces with degenerate metric are defined. It is shown that their Cartan matrices and reflection groups are affine. Due to the geometric consideration the root system structure of affine algebras is determined by a sufficiently simple algorithm.

  12. Loop realizations of quantum affine algebras

    SciTech Connect

    Cautis, Sabin; Licata, Anthony

    2012-12-15

    We give a simplified description of quantum affine algebras in their loop presentation. This description is related to Drinfeld's new realization via halves of vertex operators. We also define an idempotent version of the quantum affine algebra which is suitable for categorification.

  13. Neuroprotective effects of high affinity sigma 1 receptor selective compounds

    PubMed Central

    Luedtke, Robert R.; Perez, Evelyn; Yang, Shao-Hua; Liu, Ran; Vangveravong, Suwanna; Tu, Zhude; Mach, Robert H.; Simpkins, James W.

    2014-01-01

    We previously reported that the antipsychotic drug haloperidol, a multifunctional D2-like dopamine and sigma receptor subtype antagonist, has neuroprotective properties. In this study we further examined the association between neuroprotection and receptor antagonism by evaluating a panel of novel compounds with varying affinity at sigma and D2-like dopamine receptors. These compounds were evaluated using an in vitro cytotoxicity assay that utilizes a hippocampal-derived cell line, HT-22, in the presence or absence of varying concentrations (5 to 20 mM) of glutamate. While haloperidol was found to be a potent neuroprotective agent in this in vitro cell assay, the prototypic sigma 1 receptor agonist (+)-pentazocine was found not to be neuroprotective. Subsequently, the potency for the neuroprotection of HT-22 cells was evaluated for a) three SV series indoles which have nMolar affinity at D2-like receptors but varying affinity at sigma 1 receptor and b) two benzyl phenylacetamides sigma 1 receptor selective compounds which bind with low affinity at D2-like receptors but have nMolar affinity for the sigma 1 receptor. We observed that cytoprotection correlated with the affinity of the compounds for sigma 1 receptors. Based upon results from the HT-22 cell-based in vitro assay, two phenylacetamides, LS-127 and LS-137, were further evaluated in vivo using a transient middle cerebral artery occlusion (t-MCAO) model of stroke. At a dose of 100 µg/kg, both LS-127 and LS-137 attenuated infarct volume by approximately 50%. These studies provide further evidence that sigma 1 receptor selective compounds can provide neuroprotection in cytotoxic situations. These results also demonstrate that sigma 1 receptor selective benzyl phenylacetamides are candidate pharmacotherapeutic agents that could be used to minimize neuronal death after a stroke or head trauma. PMID:22285434

  14. Improving image segmentation by learning region affinities

    SciTech Connect

    Prasad, Lakshman; Yang, Xingwei; Latecki, Longin J

    2010-11-03

    We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

  15. Ligand Affinities Estimated by Quantum Chemical Calculations.

    PubMed

    Söderhjelm, Pär; Kongsted, Jacob; Ryde, Ulf

    2010-05-11

    We present quantum chemical estimates of ligand-binding affinities performed, for the first time, at a level of theory for which there is a hope that dispersion and polarization effects are properly accounted for (MP2/cc-pVTZ) and at the same time effects of solvation, entropy, and sampling are included. We have studied the binding of seven biotin analogues to the avidin tetramer. The calculations have been performed by the recently developed PMISP approach (polarizable multipole interactions with supermolecular pairs), which treats electrostatic interactions by multipoles up to quadrupoles, induction by anisotropic polarizabilities, and nonclassical interactions (dispersion, exchange repulsion, etc.) by explicit quantum chemical calculations, using a fragmentation approach, except for long-range interactions that are treated by standard molecular-mechanics Lennard-Jones terms. In order to include effects of sampling, 10 snapshots from a molecular dynamics simulation are studied for each biotin analogue. Solvation energies are estimated by the polarized continuum model (PCM), coupled to the multipole-polarizability model. Entropy effects are estimated from vibrational frequencies, calculated at the molecular mechanics level. We encounter several problems, not previously discussed, illustrating that we are first to apply such a method. For example, the PCM model is, in the present implementation, questionable for large molecules, owing to the use of a surface definition that gives numerous small cavities in a protein. PMID:26615702

  16. Solutions for the Mikhailov-Shabat-Yamilov Difference-Differential Equations and Generalized Solutions for the Volterra and the Toda Lattice Equations

    NASA Astrophysics Data System (ADS)

    Narita, K.

    1998-03-01

    We present two types of mixed 1-rational N-soliton solutions and two types of special solutions for four types of Volterra-related difference-differential equations arising in Mikhailov, Shabat and Yamilov's lists. We also find new expressions of mixed 1-rational N-soliton solutions for the Volterra and the Toda lattice equations based on the invariance of Gibbon and Tabor's equation (J. Math. Phys. 26 (1985), 1956) under the fractional linear transformation. By taking appropriate limits of wave numbers, we find some new rational solutions for the Volterra and the Toda lattice equations. We also present elliptic function solutions for the Volterra and the Toda lattice equations different from known ones based on the same formulation.

  17. Some introductory formalizations on the affine Hilbert spaces model of the origin of life. I. On quantum mechanical measurement and the origin of the genetic code: a general physical framework theory.

    PubMed

    Balázs, András

    2006-08-01

    A physical (affine Hilbert spaces) frame is developed for the discussion of the interdependence of the problem of the origin (symbolic assignment) of the genetic code and a possible endophysical (a kind of "internal") quantum measurement in an explicite way, following the general considerations of Balázs (Balázs, A., 2003. BioSystems 70, 43-54; Balázs, A., 2004a. BioSystems 73, 1-11). Using the Everett (a dynamic) interpretation of quantum mechanics, both the individual code assignment and the concatenated linear symbolism is discussed. It is concluded that there arises a skewed quantal probability field, with a natural dynamic non-linearity in codon assignment within the physical model adopted (essentially corresponding to a much discussed biochemical frame of self-catalyzed binding (charging) of t RNA like proto RNAs (ribozymes) with amino acids). This dynamic specific molecular complex assumption of individual code assignment, and the divergence of the code in relation to symbol concatenation, are discussed: our frame supports the former and interpret the latter as single-type codon (triplet), also unambiguous and extended assignment, selection in molecular evolution, corresponding to converging towards the fixedpoint of the internal dynamics of measurement, either in a protein- or RNA-world. In this respect, the general physical consequence is the introduction of a fourth rank semidiagonal energy tensor (see also Part II) ruling the internal dynamics as a non-linear in principle second-order one. It is inferred, as a summary, that if the problem under discussion could be expressed by the concepts of the Copenhagen interpretation of quantum mechanics in some yet not quite specified way, the matter would be particularly interesting with respect to both the origin of life and quantum mechanics, as a dynamically supported natural measurement-theoretical split between matter ("hardware") and (internal) symbolism ("software") aspects of living matter. PMID

  18. Importance of non-affine viscoelastic response in disordered fibre networks.

    PubMed

    Rizzi, L G; Auer, S; Head, D A

    2016-05-11

    Disordered fibre networks are ubiquitous in nature and have a wide range of industrial applications as novel biomaterials. Predicting their viscoelastic response is straightforward for affine deformations that are uniform over all length scales, but when affinity fails, as has been observed experimentally, modelling becomes challenging. Here we present a numerical methodology, related to an existing framework for amorphous packings, to predict the steady-state viscoelastic spectra and degree of affinity for disordered fibre networks driven at arbitrary frequencies. Applying this method to a peptide gel model reveals a monotonic increase of the shear modulus as the soft, non-affine normal modes are successively suppressed as the driving frequency increases. In addition to being dominated by fibril bending, these low frequency network modes are also shown to be delocalised. The presented methodology provides insights into the importance of non-affinity in the viscoelastic response of peptide gels, and is easily extendible to all types of fibre networks. PMID:27079274

  19. Computational design of the affinity and specificity of a therapeutic T cell receptor.

    PubMed

    Pierce, Brian G; Hellman, Lance M; Hossain, Moushumi; Singh, Nishant K; Vander Kooi, Craig W; Weng, Zhiping; Baker, Brian M

    2014-02-01

    T cell receptors (TCRs) are key to antigen-specific immunity and are increasingly being explored as therapeutics, most visibly in cancer immunotherapy. As TCRs typically possess only low-to-moderate affinity for their peptide/MHC (pMHC) ligands, there is a recognized need to develop affinity-enhanced TCR variants. Previous in vitro engineering efforts have yielded remarkable improvements in TCR affinity, yet concerns exist about the maintenance of peptide specificity and the biological impacts of ultra-high affinity. As opposed to in vitro engineering, computational design can directly address these issues, in theory permitting the rational control of peptide specificity together with relatively controlled increments in affinity. Here we explored the efficacy of computational design with the clinically relevant TCR DMF5, which recognizes nonameric and decameric epitopes from the melanoma-associated Melan-A/MART-1 protein presented by the class I MHC HLA-A2. We tested multiple mutations selected by flexible and rigid modeling protocols, assessed impacts on affinity and specificity, and utilized the data to examine and improve algorithmic performance. We identified multiple mutations that improved binding affinity, and characterized the structure, affinity, and binding kinetics of a previously reported double mutant that exhibits an impressive 400-fold affinity improvement for the decameric pMHC ligand without detectable binding to non-cognate ligands. The structure of this high affinity mutant indicated very little conformational consequences and emphasized the high fidelity of our modeling procedure. Overall, our work showcases the capability of computational design to generate TCRs with improved pMHC affinities while explicitly accounting for peptide specificity, as well as its potential for generating TCRs with customized antigen targeting capabilities. PMID:24550723

  20. Line operators in theories of class {S} , quantized moduli space of flat connections, and Toda field theory

    NASA Astrophysics Data System (ADS)

    Coman, Ioana; Gabella, Maxime; Teschner, Jörg

    2015-10-01

    Non-perturbative aspects of N=2 supersymmetric gauge theories of class S are deeply encoded in the algebra of functions on the moduli space {M}_{flat} of flat SL( N )- connections on Riemann surfaces. Expectation values of Wilson and 't Hooft line operators are related to holonomies of flat connections, and expectation values of line operators in the low-energy effective theory are related to Fock-Goncharov coordinates on {M}_{flat} . Via the decomposition of UV line operators into IR line operators, we determine their noncommutative algebra from the quantization of Fock-Goncharov Laurent polynomials, and find that it coincides with the skein algebra studied in the context of Chern-Simons theory. Another realization of the skein algebra is generated by Verlinde network operators in Toda field theory. Comparing the spectra of these two realizations provides non-trivial support for their equivalence. Our results can be viewed as evidence for the generalization of the AGT correspondence to higher-rank class S theories.

  1. Affinity Proteomics in the mountains: Alpbach 2015.

    PubMed

    Taussig, Michael J

    2016-09-25

    The 2015 Alpbach Workshop on Affinity Proteomics, organised by the EU AFFINOMICS consortium, was the 7th workshop in this series. As in previous years, the focus of the event was the current state of affinity methods for proteome analysis, including complementarity with mass spectrometry, progress in recombinant binder production methods, alternatives to classical antibodies as affinity reagents, analysis of proteome targets, industry focus on biomarkers, and diagnostic and clinical applications. The combination of excellent science with Austrian mountain scenery and winter sports engender an atmosphere that makes this series of workshops exceptional. The articles in this Special Issue represent a cross-section of the presentations at the 2015 meeting. PMID:27118167

  2. Optimized Affinity Capture of Yeast Protein Complexes.

    PubMed

    LaCava, John; Fernandez-Martinez, Javier; Hakhverdyan, Zhanna; Rout, Michael P

    2016-01-01

    Here, we describe an affinity isolation protocol. It uses cryomilled yeast cell powder for producing cell extracts and antibody-conjugated paramagnetic beads for affinity capture. Guidelines for determining the optimal extraction solvent composition are provided. Captured proteins are eluted in a denaturing solvent (sodium dodecyl sulfate polyacrylamide gel electrophoresis sample buffer) for gel-based proteomic analyses. Although the procedures can be modified to use other sources of cell extract and other forms of affinity media, to date we have consistently obtained the best results with the method presented. PMID:27371596

  3. Aptamers in Affinity Separations: Stationary Separation

    NASA Astrophysics Data System (ADS)

    Ravelet, Corinne; Peyrin, Eric

    The use of DNA or RNA aptamers as tools in analytical chemistry is a very promising field of research because of their capabilities to bind specifically the target molecules with an affinity similar to that of antibodies. Notably, they appear to be of great interest as target-specific ligands for the separation and capture of various analytes in affinity chromatography and related affinity-based methods such as magnetic bead technology. In this chapter, the recent developments of these aptamer-based separation/capture approaches are addressed.

  4. Affinity purification of heme-tagged proteins.

    PubMed

    Asher, Wesley B; Bren, Kara L

    2014-01-01

    Protein affinity purification techniques are widely used for isolating pure target proteins for biochemical and structural characterization. Herein, we describe the protocol for affinity-based purification of proteins expressed in Escherichia coli that uses the coordination of a peptide tag covalently modified with heme c, known as a heme-tag, to an L-histidine immobilized Sepharose resin. This approach provides an affinity purification tag visible to the eye, facilitating tracking of the protein. In addition, we describe methods for specifically detecting heme-tagged proteins in SDS-PAGE gels using a heme-staining procedure and for quantifying the proteins using a pyridine hemochrome assay. PMID:24943311

  5. A molecular determinant of phosphoinositide affinity in mammalian TRPV channels

    PubMed Central

    Velisetty, Phanindra; Borbiro, Istvan; Kasimova, Marina A.; Liu, Luyu; Badheka, Doreen; Carnevale, Vincenzo; Rohacs, Tibor

    2016-01-01

    Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] is an important cofactor for ion channels. Affinity for this lipid is a major determinant of channel inhibition by depletion of PI(4,5)P2 upon phospholipase C (PLC) activation. Little is known about what determines PI(4,5)P2 affinity in mammalian ion channels. Here we report that two members of the Transient Receptor Potential Vanilloid (TRPV) ion channel family, TRPV5 and TRPV6 lack a positively charged residue in the TM4-TM5 loop that was shown to interact with PI(4,5)P2 in TRPV1, which shows high affinity for this lipid. When this positively charged residue was introduced to either TRPV6 or TRPV5, they displayed markedly higher affinities for PI(4,5)P2, and were largely resistant to inhibition by PI(4,5)P2 depletion. Furthermore, Ca2+-induced inactivation of TRPV6 was essentially eliminated in the G488R mutant, showing the importance of PLC-mediated PI(4,5)P2 depletion in this process. Computational modeling shows that the introduced positive charge interacts with PI(4,5)P2 in TRPV6. PMID:27291418

  6. Chemokines and the Signaling Modules Regulating Integrin Affinity

    PubMed Central

    Montresor, Alessio; Toffali, Lara; Constantin, Gabriela; Laudanna, Carlo

    2012-01-01

    Integrin-mediated adhesion is a general concept referring to a series of adhesive phenomena including tethering–rolling, affinity, valency, and binding stabilization altogether controlling cell avidity (adhesiveness) for the substrate. Arrest chemokines modulate each aspect of integrin activation, although integrin affinity regulation has been recognized as the prominent event in rapid leukocyte arrest induced by chemokines. A variety of inside-out and outside-in signaling mechanisms have been related to the process of integrin-mediated adhesion in different cellular models, but only few of them have been clearly contextualized to rapid integrin affinity modulation by arrest chemokines in primary leukocytes. Complex signaling processes triggered by arrest chemokines and controlling leukocyte integrin activation have been described for ras-related rap and for rho-related small GTPases. We summarize the role of rap and rho small GTPases in the regulation of rapid integrin affinity in primary leukocytes and provide a modular view of these pro-adhesive signaling events. A potential, albeit still speculative, mechanism of rho-mediated regulation of cytoskeletal proteins controlling the last step of integrin activation is also discussed. We also discuss data suggesting a functional integration between the rho- and rap-modules of integrin activation. Finally we examine the universality of signaling mechanisms regulating integrin triggering by arrest chemokines. PMID:22654882

  7. Affinity and Avidity in Antibody-Based Tumor Targeting

    PubMed Central

    Rudnick, Stephen I.

    2009-01-01

    Summation Many factors contribute to successful tumor targeting by antibodies. Besides properties of the tumor tissue and general antibody pharmacology, a relationship exists between an antibody and its antigen that can shape penetration, catabolism, specificity, and efficacy. The affinity and avidity of the binding interactions play critical roles in these dynamics. In this work, we review the principles that guide models predicting tumor penetration and cellular internalization while providing a critical overview of studies aimed at experimentally determining the specific role of affinity and avidity in these processes. One should gain the perspective that binding affinity can, in part, dictate the localization of antibodies in tumors, leading to high concentrations in the perivascular space or low concentrations diffused throughout the tumor. These patterns can be simply due to the diminution of available dose by binding antigen and are complicated by internalization and degradation stemming from slow rates of dissociation. As opposed to the trend of simply increasing affinity to increase efficacy, novel strategies that increase avidity and broaden specificity have made significant progress in tumor targeting. PMID:19409036

  8. A molecular determinant of phosphoinositide affinity in mammalian TRPV channels.

    PubMed

    Velisetty, Phanindra; Borbiro, Istvan; Kasimova, Marina A; Liu, Luyu; Badheka, Doreen; Carnevale, Vincenzo; Rohacs, Tibor

    2016-01-01

    Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] is an important cofactor for ion channels. Affinity for this lipid is a major determinant of channel inhibition by depletion of PI(4,5)P2 upon phospholipase C (PLC) activation. Little is known about what determines PI(4,5)P2 affinity in mammalian ion channels. Here we report that two members of the Transient Receptor Potential Vanilloid (TRPV) ion channel family, TRPV5 and TRPV6 lack a positively charged residue in the TM4-TM5 loop that was shown to interact with PI(4,5)P2 in TRPV1, which shows high affinity for this lipid. When this positively charged residue was introduced to either TRPV6 or TRPV5, they displayed markedly higher affinities for PI(4,5)P2, and were largely resistant to inhibition by PI(4,5)P2 depletion. Furthermore, Ca(2+)-induced inactivation of TRPV6 was essentially eliminated in the G488R mutant, showing the importance of PLC-mediated PI(4,5)P2 depletion in this process. Computational modeling shows that the introduced positive charge interacts with PI(4,5)P2 in TRPV6. PMID:27291418

  9. PRINCIPLES OF AFFINITY-BASED BIOSENSORS

    EPA Science Inventory

    Despite the amount of resources that have been invested by national and international academic, government, and commercial sectors to develop affinity-based biosensor products, little obvious success has been realized through commercialization of these devices for specific applic...

  10. Minimal information to determine affine shape equivalence.

    PubMed

    Wagemans, J; Van Gool, L; Lamote, C; Foster, D H

    2000-04-01

    Participants judged the affine equivalence of 2 simultaneously presented 4-point patterns. Performance level (d') varied between 1.5 and 2.7, depending on the information available for solving the correspondence problem (insufficient in Experiment 1a, superfluous in Experiment 1b, and minimal in Experiments 1c, 2a, 2b) and on the exposure time (unlimited in Experiments 1 and 2a and 500 ms in Experiment 2b), but it did not vary much with the complexity of the affine transformation (rotation and slant in Experiment 1 and same plus tilt in Experiment 2). Performance in Experiment 3 was lower with 3-point patterns than with 4-point patterns, whereas blocking the trials according to the affine transformation parameters had little effect. Determining affine shape equivalence with minimal-information displays is based on a fast assessment of qualitatively or quasi-invariant properties such as convexity/ concavity, parallelism, and collinearity. PMID:10811156

  11. Protein purification using PDZ affinity chromatography.

    PubMed

    Walkup, Ward G; Kennedy, Mary B

    2015-01-01

    PDZ domains function in nature as protein-binding domains within scaffold and membrane-associated proteins. They comprise approximately 90 residues and undergo specific, high-affinity interactions with complementary C-terminal peptide sequences, other PDZ domains, and/or phospholipids. We have previously shown that the specific, strong interactions of PDZ domains with their ligands make them well suited for use in affinity chromatography. This unit provides protocols for the PDZ affinity chromatography procedure that are applicable for the purification of proteins that contain PDZ domains or PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We detail the preparation of affinity resins composed of PDZ domains or PDZ domain peptide ligands coupled to solid supports. These resins can be used to purify proteins containing endogenous or genetically introduced PDZ domains or ligands, eluting the proteins with free PDZ domain peptide ligands. PMID:25829303

  12. Visualizing antibody affinity maturation in germinal centers.

    PubMed

    Tas, Jeroen M J; Mesin, Luka; Pasqual, Giulia; Targ, Sasha; Jacobsen, Johanne T; Mano, Yasuko M; Chen, Casie S; Weill, Jean-Claude; Reynaud, Claude-Agnès; Browne, Edward P; Meyer-Hermann, Michael; Victora, Gabriel D

    2016-03-01

    Antibodies somatically mutate to attain high affinity in germinal centers (GCs). There, competition between B cell clones and among somatic mutants of each clone drives an increase in average affinity across the population. The extent to which higher-affinity cells eliminating competitors restricts clonal diversity is unknown. By combining multiphoton microscopy and sequencing, we show that tens to hundreds of distinct B cell clones seed each GC and that GCs lose clonal diversity at widely disparate rates. Furthermore, efficient affinity maturation can occur in the absence of homogenizing selection, ensuring that many clones can mature in parallel within the same GC. Our findings have implications for development of vaccines in which antibodies with nonimmunodominant specificities must be elicited, as is the case for HIV-1 and influenza. PMID:26912368

  13. Nanoparticle Surface Affinity as a Predictor of Trophic Transfer.

    PubMed

    Geitner, Nicholas K; Marinakos, Stella M; Guo, Charles; O'Brien, Niall; Wiesner, Mark R

    2016-07-01

    Nanoscale materials, whether natural, engineered, or incidental, are increasingly acknowledged as important components in large, environmental systems with potential implications for environmental impact and human health. Mathematical models are a useful tool for handling the rapidly increasing complexity and diversity of these materials and their exposure routes. Presented here is a mathematical model of trophic transfer driven by nanomaterial surface affinity for environmental and biological surfaces, developed in tandem with an experimental functional assay for determining these surface affinities. We found that nanoparticle surface affinity is a strong predictor of uptake through predation in a simple food web consisting of the algae Chlorella vulgaris and daphnid Daphnia magna. The mass of nanoparticles internalized by D. magna through consuming nanomaterial-contaminated algae varied linearly with surface-attachment efficiency. Internalized quantities of gold nanoparticles in D. magna ranged from 8.3 to 23.6 ng/mg for nanoparticle preparations with surface-attachment efficiencies ranging from 0.07 to 1. This model, coupled with the functional-assay approach, may provide a useful screening tool for existing materials as well as a predictive model for their development. PMID:27249534

  14. Rapid D-Affine Biventricular Cardiac Function with Polar Prediction

    PubMed Central

    Gilbert, Kathleen; Cowan, Brett; Suinesiaputra, Avan; Occleshaw, Christopher; Young, Alistair

    2014-01-01

    Although many solutions have been proposed for left ventricular functional analysis of the heart, right and left (bi-) ventricular function has been problematic due to the complex geometry and large motions. Biventricular function is particularly important in congenital heart disease, the most common type of birth defects. We describe a rapid interactive analysis tool for biventricular function which incorporates 1) a 3D+ time finite element model of biventricular geometry, 2) a fast prediction step which estimates an initial geometry in a polar coordinate system, and 3) a Cartesian update which penalizes deviations from affine transformations (D-Affine) from a prior. Solution times were very rapid, enabling interaction in real time using guide point modeling. The method was applied to 13 patients with congenital heart disease and compared with the clinical gold standard of manual tracing. Results between the methods showed good correlation (R2 > 0.9) and good precision (volume<17ml; mass<11g) for both chambers. PMID:25485422

  15. Affinity engineering of maltoporin: variants with enhanced affinity for particular ligands.

    PubMed

    Clune, A; Lee, K S; Ferenci, T

    1984-05-31

    Affinity-chromatographic selection on immobilized starch was used to selectively enhance the affinity of the maltodextrin-specific pore protein ( maltoporin , LamB protein, or lambda receptor protein) in the outer membrane of E. coli. Selection strategies were established for rare bacteria in large populations producing maltoporin variants with enhanced affinities for both starch and maltose, for starch but not maltose and for maltose but not starch. Three classes of lamB mutants with up to eight-fold increase in affinity for particular ligands were isolated. These mutants provide a unique range of modifications in the specificity of a transport protein. PMID:6375667

  16. Classification of neocortical interneurons using affinity propagation

    PubMed Central

    Santana, Roberto; McGarry, Laura M.; Bielza, Concha; Larrañaga, Pedro; Yuste, Rafael

    2013-01-01

    In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. In fact, neuronal classification is a difficult problem because it is unclear how to designate a neuronal cell class and what are the best characteristics to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological, or molecular characteristics, have provided quantitative and unbiased identification of distinct neuronal subtypes, when applied to selected datasets. However, better and more robust classification methods are needed for increasingly complex and larger datasets. Here, we explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. Affinity propagation outperformed Ward's method, a current standard clustering approach, in classifying the neurons into 4 subtypes. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits. PMID:24348339

  17. Classification of neocortical interneurons using affinity propagation.

    PubMed

    Santana, Roberto; McGarry, Laura M; Bielza, Concha; Larrañaga, Pedro; Yuste, Rafael

    2013-01-01

    In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. In fact, neuronal classification is a difficult problem because it is unclear how to designate a neuronal cell class and what are the best characteristics to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological, or molecular characteristics, have provided quantitative and unbiased identification of distinct neuronal subtypes, when applied to selected datasets. However, better and more robust classification methods are needed for increasingly complex and larger datasets. Here, we explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. Affinity propagation outperformed Ward's method, a current standard clustering approach, in classifying the neurons into 4 subtypes. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits. PMID:24348339

  18. Affinity purification of aprotinin from bovine lung.

    PubMed

    Xin, Yu; Liu, Lanhua; Chen, Beizhan; Zhang, Ling; Tong, Yanjun

    2015-05-01

    An affinity protocol for the purification of aprotinin from bovine lung was developed. To simulate the structure of sucrose octasulfate, a natural specific probe for aprotinin, the affinity ligand was composed of an acidic head and a hydrophobic stick, and was then linked with Sepharose. The sorbent was then subjected to adsorption analysis with pure aprotinin. The purification process consisted of one step of affinity chromatography and another step of ultrafiltration. Then purified aprotinin was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, trypsin inhibitor activity, gel-filtration, and thin-layer chromatography analysis. As calculated, the theoretical maximum adsorption (Qmax ) of the affinity sorbent was 25,476.0 ± 184.8 kallikrein inactivator unit/g wet gel; the dissociation constant of the complex "immobilized ligand-aprotinin" (Kd ) was 4.6 ± 0.1 kallikrein inactivator unit/mL. After the affinity separation of bovine lung aprotinin, reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis and gel-filtration chromatography revealed that the protein was a single polypeptide, and the purities were ∼ 97 and 100%, respectively; the purified peptide was also confirmed with aprotinin standard by gel-filtration chromatography and thin-layer chromatography. After the whole purification process, protein, and bioactivity recoveries were 2.2 and 92.6%, respectively; and the specific activity was up to 15,907.1 ± 10.2 kallikrein inactivator unit/mg. PMID:25677462

  19. Identity, Affinity, Reality: Making the Case for Affinity Groups in Elementary School

    ERIC Educational Resources Information Center

    Parsons, Julie; Ridley, Kimberly

    2012-01-01

    Affinity groups are places where students build connections and process "ouch" moments from their classes. Children talk about the isolation they sometimes feel. The relationships students gain through race-based affinity groups enable them to feel less alone with their emotions and help them build a stronger sense of self. At the same time,…

  20. The dynamics of metric-affine gravity

    SciTech Connect

    Vitagliano, Vincenzo; Sotiriou, Thomas P.; Liberati, Stefano

    2011-05-15

    Highlights: > The role and the dynamics of the connection in metric-affine theories is explored. > The most general second order action does not lead to a dynamical connection. > Including higher order invariants excites new degrees of freedom in the connection. > f(R) actions are also discussed and shown to be a non- representative class. - Abstract: Metric-affine theories of gravity provide an interesting alternative to general relativity: in such an approach, the metric and the affine (not necessarily symmetric) connection are independent quantities. Furthermore, the action should include covariant derivatives of the matter fields, with the covariant derivative naturally defined using the independent connection. As a result, in metric-affine theories a direct coupling involving matter and connection is also present. The role and the dynamics of the connection in such theories is explored. We employ power counting in order to construct the action and search for the minimal requirements it should satisfy for the connection to be dynamical. We find that for the most general action containing lower order invariants of the curvature and the torsion the independent connection does not carry any dynamics. It actually reduces to the role of an auxiliary field and can be completely eliminated algebraically in favour of the metric and the matter field, introducing extra interactions with respect to general relativity. However, we also show that including higher order terms in the action radically changes this picture and excites new degrees of freedom in the connection, making it (or parts of it) dynamical. Constructing actions that constitute exceptions to this rule requires significant fine tuned and/or extra a priori constraints on the connection. We also consider f(R) actions as a particular example in order to show that they constitute a distinct class of metric-affine theories with special properties, and as such they cannot be used as representative toy theories to

  1. Displacement phenomena in lectin affinity chromatography.

    PubMed

    Cho, Wonryeon

    2015-10-01

    The work described here examines displacement phenomena that play a role in lectin affinity chromatography and their potential to impact reproducibility. This was achieved using Lycopersicon esculentum lectin (LEL), a lectin widely used in monitoring cancer. Four small identical LEL columns were coupled in series to form a single affinity chromatography system with the last in the series connected to an absorbance detector. The serial affinity column set (SACS) was then loaded with human plasma proteins. At the completion of loading, the column set was disassembled, the four columns were eluted individually, the captured proteins were trypsin digested, the peptides were deglycosylated with PNGase F, and the parent proteins were identified through mass spectral analyses. Significantly different sets of glycoproteins were selected by each column, some proteins appearing to be exclusively bound to the first column while others were bound further along in the series. Clearly, sample displacement chromatography (SDC) occurs. Glycoproteins were bound at different places in the column train, identifying the presence of glycoforms with different affinity on a single glycoprotein. It is not possible to see these phenomena in the single column mode of chromatography. Moreover, low abundance proteins were enriched, which facilitates detection. The great advantage of this method is that it differentiates between glycoproteins on the basis of their binding affinity. Displacement phenomena are concluded to be a significant component of the separation mechanism in heavily loaded lectin affinity chromatography columns. This further suggests that care must be exercised in sample loading of lectin columns to prevent analyte displacement with nonretained proteins. PMID:26348026

  2. Negative Electron Affinity Mechanism for Diamond Surfaces

    NASA Technical Reports Server (NTRS)

    Krainsky, I. L.; Asnin, V. M.

    1998-01-01

    The energy distribution of the secondary electrons for chemical vacuum deposited diamond films with Negative Electron Affinity (NEA) was investigated. It was found that while for completely hydrogenated diamond surfaces the negative electron affinity peak in the energy spectrum of the secondary electrons is present for any energy of the primary electrons, for partially hydrogenated diamond surfaces there is a critical energy above which the peak is present in the spectrum. This critical energy increases sharply when hydrogen coverage of the diamond surface diminishes. This effect was explained by the change of the NEA from the true type for the completely hydrogenated surface to the effective type for the partially hydrogenated surfaces.

  3. New unitary affine-Virasoro constructions

    SciTech Connect

    Halpern, M.B.; Kiritsis, E.; Obers, N.A.; Poratti, M. ); Yamron, J.P. )

    1990-06-20

    This paper reports on a quasi-systematic investigation of the Virasoro master equation. The space of all affine-Virasoro constructions is organized by K-conjugation into affine-Virasoro nests, and an estimate of the dimension of the space shows that most solutions await discovery. With consistent ansatze for the master equation, large classes of new unitary nests are constructed, including quadratic deformation nests with continuous conformal weights, and unitary irrational central charge nests, which may dominate unitary rational central charge on compact g.

  4. Adsorption affinity of anions on metal oxyhydroxides

    NASA Astrophysics Data System (ADS)

    Pechenyuk, S. I.; Semushina, Yu. P.; Kuz'mich, L. F.

    2013-03-01

    The dependences of anion (phosphate, carbonate, sulfate, chromate, oxalate, tartrate, and citrate) adsorption affinity anions from geometric characteristics, acid-base properties, and complex forming ability are generalized. It is shown that adsorption depends on the nature of both the anions and the ionic medium and adsorbent. It is established that anions are generally grouped into the following series of adsorption affinity reduction: PO{4/3-}, CO{3/2-} > C2O{4/2-}, C(OH)(CH2)2(COO){3/3-}, (CHOH)2(COO){2/2-} > CrO{4/2-} ≫ SO{4/2-}.

  5. Structure of Greyhound hemoglobin: origin of high oxygen affinity.

    PubMed

    Bhatt, Veer S; Zaldívar-López, Sara; Harris, David R; Couto, C Guillermo; Wang, Peng G; Palmer, Andre F

    2011-05-01

    This study presents the crystal structure of Greyhound hemoglobin (GrHb) determined to 1.9 Å resolution. GrHb was found to crystallize with an α₁β₁ dimer in the asymmetric unit and belongs to the R2 state. Oxygen-affinity measurements combined with the fact that GrHb crystallizes in the R2 state despite the high-salt conditions used for crystallization strongly indicate that GrHb can serve as a model high-oxygen-affinity hemoglobin (Hb) for higher mammals, especially humans. Structural analysis of GrHb and its comparison with the R2-state of human Hb revealed several regions that can potentially contribute to the high oxygen affinity of GrHb and serve to rationalize the additional stability of the R2-state of GrHb. A previously well studied hydrophobic cluster of bar-headed goose Hb near α119 was also incorporated in the comparison between GrHb and human Hb. Finally, a structural comparison with generic dog Hb and maned wolf Hb was conducted, revealing that in contrast to GrHb these structures belong to the R state of Hb and raising the intriguing possibility of an additional allosteric factor co-purifying with GrHb that can modulate its quaternary structure. PMID:21543841

  6. Affinity labeling of the ribosomal P site in Drosophila melanogaster

    SciTech Connect

    North, D.

    1987-01-01

    Several recent studies have probed the peptidyl transferase region of the Drosophila ribosome via the use of reactive site specific analogues (affinity labels). P site proteins adjacent to the 3' end of the amino acid bearing tRNA strand were labeled with modified tRNA fragments. Drugs affecting the binding of these agents were used to further clarify the nature of the region. The nascent peptide region of the P site was not labeled in previous experiments. To label that region radioactive Bromoacetylphenylalanyl-tRNA (BrAcphe-tRNA) was synthesized. The alpha-bromoacetyl group of this analogue is potentially reactive with nucleophiles present in either proteins or RNAs. Charged tRNAs and tRNA analogues bearing a peptide bond on the N-terminus of their amino acid are recognized as having affinity for the ribosomal P site. Specific labeling of the P site by BrAcphe-tRNA was confirmed by its ability to radioactively label proteins indirectly. As many as 8 ribosomal proteins may be labeled under these conditions, however, the majority of the bound label is associated with 3 large subunit proteins and 2 small subunit proteins. Overlaps between the proteins labeled by BrAcphe-tRNA and those labeled by other affinity labels are examined and a model of the peptidyl transferase region of Drosophila ribosomes is presented.

  7. Controlling Affinity Binding with Peptide-Functionalized Poly(ethylene glycol) Hydrogels**

    PubMed Central

    Lin, Chien-Chi; Anseth, Kristi S.

    2009-01-01

    Poly(ethylene glycol) (PEG) hydrogels functionalized with peptide moieties have been widely used in regenerative medicine applications. While many studies have suggested the importance of affinity binding within PEG hydrogels, the relationships between the structures of the peptide motifs and their binding to protein therapeutics remain largely unexplored, especially in the recently developed thiol-acrylate photopolymerization systems. Herein, we employ Förster resonance energy transfer (FRET) and thiol-acrylate photopolymerizations to investigate how the architectures of affinity peptides in crosslinked hydrogels affect their binding to diffusible proteins. The binding between diffusible streptavidin and biotinylated peptide immobilized to PEG hydrogel network was used as a model system to reveal the interplay between affinity binding and peptide sequences/architectures. In addition, we design peptides with different structures to enhance affinity binding within PEG hydrogels and to provide tunable affinity-based controlled delivery of basic fibroblast growth factor (bFGF). This study demonstrates the importance of affinity binding in controlling the availability of hydrogel-encapsulated proteins and provides strategies for enhancing affinity binding of protein therapeutics to bound peptide moieties in thiol-acrylate photopolymerized PEG hydrogels. The results presented herein should find useful on the design and fabrication of hydrogels to retain and sustained release of growth factors for promoting tissue regeneration. PMID:20148198

  8. Negative Electron Affinity Effect on the Surface of Chemical Vapor Deposited Diamond Polycrystalline Films

    NASA Technical Reports Server (NTRS)

    Krainsky, I. L.; Asnin, V. M.; Mearini, G. T.; Dayton, J. A., Jr.

    1996-01-01

    Strong negative electron affinity effects have been observed on the surface of as-grown chemical vapor deposited diamond using Secondary Electron Emission. The test samples were randomly oriented and the surface was terminated with hydrogen. The effect appears as an intensive peak in the low energy part of the spectrum of the electron energy distribution and may be described in the model of effective negative electron affinity.

  9. Streamlining the Pipeline for Generation of Recombinant Affinity Reagents by Integrating the Affinity Maturation Step.

    PubMed

    Huang, Renhua; Gorman, Kevin T; Vinci, Chris R; Dobrovetsky, Elena; Gräslund, Susanne; Kay, Brian K

    2015-01-01

    Often when generating recombinant affinity reagents to a target, one singles out an individual binder, constructs a secondary library of variants, and affinity selects a tighter or more specific binder. To enhance the throughput of this general approach, we have developed a more integrated strategy where the "affinity maturation" step is part of the phage-display pipeline, rather than a follow-on process. In our new schema, we perform two rounds of affinity selection, followed by error-prone PCR on the pools of recovered clones, generation of secondary libraries, and three additional rounds of affinity selection, under conditions of off-rate competition. We demonstrate the utility of this approach by generating low nanomolar fibronectin type III (FN3) monobodies to five human proteins: ubiquitin-conjugating enzyme E2 R1 (CDC34), COP9 signalosome complex subunit 5 (COPS5), mitogen-activated protein kinase kinase 5 (MAP2K5), Splicing factor 3A subunit 1 (SF3A1) and ubiquitin carboxyl-terminal hydrolase 11 (USP11). The affinities of the resulting monobodies are typically in the single-digit nanomolar range. We demonstrate the utility of two binders by pulling down the targets from a spiked lysate of HeLa cells. This integrated approach should be applicable to directed evolution of any phage-displayed affinity reagent scaffold. PMID:26437402

  10. Streamlining the Pipeline for Generation of Recombinant Affinity Reagents by Integrating the Affinity Maturation Step

    PubMed Central

    Huang, Renhua; Gorman, Kevin T.; Vinci, Chris R.; Dobrovetsky, Elena; Gräslund, Susanne; Kay, Brian K.

    2015-01-01

    Often when generating recombinant affinity reagents to a target, one singles out an individual binder, constructs a secondary library of variants, and affinity selects a tighter or more specific binder. To enhance the throughput of this general approach, we have developed a more integrated strategy where the “affinity maturation” step is part of the phage-display pipeline, rather than a follow-on process. In our new schema, we perform two rounds of affinity selection, followed by error-prone PCR on the pools of recovered clones, generation of secondary libraries, and three additional rounds of affinity selection, under conditions of off-rate competition. We demonstrate the utility of this approach by generating low nanomolar fibronectin type III (FN3) monobodies to five human proteins: ubiquitin-conjugating enzyme E2 R1 (CDC34), COP9 signalosome complex subunit 5 (COPS5), mitogen-activated protein kinase kinase 5 (MAP2K5), Splicing factor 3A subunit 1 (SF3A1) and ubiquitin carboxyl-terminal hydrolase 11 (USP11). The affinities of the resulting monobodies are typically in the single-digit nanomolar range. We demonstrate the utility of two binders by pulling down the targets from a spiked lysate of HeLa cells. This integrated approach should be applicable to directed evolution of any phage-displayed affinity reagent scaffold. PMID:26437402

  11. High affinity of lead for fetal haemoglobin.

    PubMed Central

    Ong, C N; Lee, W R

    1980-01-01

    In-vitro experiments using 203Pb were performed to identify lead-binding components in human haemoglobin. Sephadex A-50 ion-exchange chromatography of haemolysate showed that different types of haemoglobin had different affinities for lead. For the haemolysate from adults, lead was present in both Hb A (alpha 2 beta 2) and Hb A2 (alpha 2 delta 2), whereas, in the haemolysate from new-born infants, the haemoglobin of fetal origin, Hb F (alpha 2 gamma 2) showed a much greater affinity for 203Pb than the adult haemoglobin Hb A (alpha 2 beta 2), obtained from maternal blood. Analysis of the 203 Pb-labelled haemoglobin suggested that about 82% of 203Pb was in the globin polypeptide. Further analysis with carboxylmethyl (CM) cellulose chromatography indicated that the gamma globin of fetal origin had a higher affinity for 203Pb than the beta globin, whereas alpha globin appeared to be unimportant in lead binding. The results of the different affinities for lead of different Hb types are discussed with regard to the effect of lead upon haemoglobin synthesis. PMID:6158989

  12. Vygotsky's and Buber's Pedagogical Perspectives: Some Affinities

    ERIC Educational Resources Information Center

    Bartholo, Roberto; Tunes, Elizabeth; Tacca, Maria Carmen Villela Rosa

    2010-01-01

    The purpose of this paper is to examine the dialogical and creative character of pedagogic work by analyzing the affinities between Martin Buber's "I-Thou relation" and Lev Semenovich Vygotsky's "Zone of Proximal Development". Backed up by empirical studies on the teacher-student relation, we understand that education can only result in students'…

  13. Affine calibration based on invariable extrinsic parameters for stereo light microscope

    NASA Astrophysics Data System (ADS)

    Li, Weixian; Wei, Zhenzhong; Zhang, Guangjun

    2014-10-01

    The stereo light microscope (SLM) plays an important role in the measurement of three-dimensional geometry on the microscopic scale. We propose a fast and precise affine calibration algorithm based on the invariable extrinsic parameters for the SLM. This calibration algorithm with a free planar reference consists of three steps: first, derive the extrinsic parameters based on their invariable definition in the pinhole and affine models; second, calculate the intrinsic parameters through homography matrix; finally, refine all the model parameters by global optimization with the previous closed-form solutions as the initial values. The effectiveness of assuming a noncoaxial optical system as an affine camera is also verified to affinely model all types of SLMs. The calibration experiments show that the affine calibration is preferable for multicriteria including running time, relative positioning precision, and absolute positioning precision. With PlanApo S 1.5× and a total magnification of 3.024×, the proposed affine calibration algorithm achieves a distance error of 0.423 μm and a positioning error of 0.195 mm within 10.6 s.

  14. Analysis of free drug fractions in human serum by ultrafast affinity extraction and two-dimensional affinity chromatography.

    PubMed

    Zheng, Xiwei; Podariu, Maria; Matsuda, Ryan; Hage, David S

    2016-01-01

    Ultrafast affinity extraction and a two-dimensional high performance affinity chromatographic system were used to measure the free fractions for various drugs in serum and at typical therapeutic concentrations. Pooled samples of normal serum or serum from diabetic patients were utilized in this work. Several drug models (i.e., quinidine, diazepam, gliclazide, tolbutamide, and acetohexamide) were examined that represented a relatively wide range of therapeutic concentrations and affinities for human serum albumin (HSA). The two-dimensional system consisted of an HSA microcolumn for the extraction of a free drug fraction, followed by a larger HSA analytical column for the further separation and measurement of this fraction. Factors that were optimized in this method included the flow rates, column sizes, and column switching times that were employed. The final extraction times used for isolating the free drug fractions were 333-665 ms or less. The dissociation rate constants for several of the drugs with soluble HSA were measured during system optimization, giving results that agreed with reference values. In the final system, free drug fractions in the range of 0.7-9.5% were measured and gave good agreement with values that were determined by ultrafiltration. Association equilibrium constants or global affinities were also estimated by this approach for the drugs with soluble HSA. The results for the two-dimensional system were obtained in 5-10 min or less and required only 1-5 μL of serum per injection. The same approach could be adapted for work with other drugs and proteins in clinical samples or for biomedical research. PMID:26462924

  15. Quantitative analysis of multiple kappa-opioid receptors by selective and nonselective ligand binding in guinea pig spinal cord: Resolution of high and low affinity states of the kappa 2 receptors by a computerized model-fitting technique

    SciTech Connect

    Tiberi, M.; Magnan, J. )

    1990-05-01

    The binding characteristics of selective and nonselective opioids have been studied in whole guinea pig spinal cord, using a computer fitting method to analyze the data obtained from saturation and competition studies. The delineation of specific binding sites labeled by the mu-selective opioid (3H)D-Ala2,MePhe4,Gly-ol5-enkephalin (Kd = 2.58 nM, R = 4.52 pmol/g of tissue) and by the delta-selective opioid (3H)D-Pen2, D-Pen5-enkephalin (Kd = 2.02 nM, R = 1.47 pmol/g of tissue) suggests the presence of mu and delta-receptors in the spinal cord tissue. The presence of kappa receptors was probed by the kappa-selective opioid (3H)U69593 (Kd = 3.31 nM, R = 2.00 pmol/g of tissue). The pharmacological characterization of the sites labeled by (3H)U69593 confirms the assumption that this ligand discriminates kappa receptors in guinea pig spinal cord. The benzomorphan (3H)ethylketazocine labels a population of receptors with one homogeneous affinity state (Kd = 0.65 nM, R = 7.39 pmol/g of tissue). The total binding capacity of this ligand was not different from the sum of the binding capacities of mu, delta-, and kappa-selective ligands. Under mu- and delta-suppressed conditions, (3H)ethylketazocine still binds to receptors with one homogeneous affinity state (Kd = 0.45 nM, R = 1.69 pmol/g of tissue). Competition studies performed against the binding of (3H)ethylketazocine under these experimental conditions reveal that the pharmacological profile of the radiolabeled receptors is similar to the profile of the kappa receptors labeled with (3H)U69593. Saturation studies using the nonselective opioid (3H)bremazocine demonstrate that this ligand binds to spinal cord membranes with heterogeneous affinities (Kd1 = 0.28 nM, R1 = 7.91 pmol/g of tissue; Kd2 = 3.24 nM, R2 = 11.2 pmol/g of tissue).

  16. Affinity purification of metalloprotease from marine bacterium using immobilized metal affinity chromatography.

    PubMed

    Li, Shangyong; Wang, Linna; Yang, Juan; Bao, Jing; Liu, Junzhong; Lin, Shengxiang; Hao, Jianhua; Sun, Mi

    2016-06-01

    In this study, an efficient affinity purification protocol for an alkaline metalloprotease from marine bacterium was developed using immobilized metal affinity chromatography. After screening and optimization of the affinity ligands and spacer arm lengths, Cu-iminmodiacetic acid was chosen as the optimal affinity ligand, which was coupled to Sepharose 6B via a 14-atom spacer arm. The absorption analysis of this medium revealed a desorption constant Kd of 21.5 μg/mL and a theoretical maximum absorption Qmax of 24.9 mg/g. Thanks to this affinity medium, the enzyme could be purified by only one affinity purification step with a purity of approximately 95% pure when analyzed by high-performance liquid chromatography and reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis. The recovery of the protease activity reached 74.6%, which is much higher than the value obtained by traditional protocols (8.9%). These results contribute to the industrial purifications and contribute a significant reference for the purification of other metalloproteases. PMID:27058973

  17. Self-affine surface morphology of plastically deformed metals.

    PubMed

    Zaiser, Michael; Grasset, Frederic Madani; Koutsos, Vasileios; Aifantis, Elias C

    2004-11-01

    We analyze the surface morphology of metals after plastic deformation over a range of scales from 10 nm to 2 mm using atomic force microscopy and scanning white-light interferometry. We demonstrate that an initially smooth surface during deformation develops self-affine roughness over almost 4 orders of magnitude in scale. The Hurst exponent H of one-dimensional surface profiles initially decreases with increasing strain and then stabilizes at H approximately 0.75. We show that the profiles can be mathematically modeled as graphs of a fractional Brownian motion. Our findings can be understood in terms of a fractal distribution of plastic strain within the deformed samples. PMID:15600851

  18. Synthesis and structure-activity relationships of a new model of arylpiperazines. Part 7: Study of the influence of lipophilic factors at the terminal amide fragment on 5-HT(1A) affinity/selectivity.

    PubMed

    López-Rodríguez, María L; Ayala, David; Viso, Alma; Benhamú, Bellinda; de La Pradilla, Roberto Fernández; Zarza, Fernando; Ramos, José A

    2004-03-15

    The influence of lipophilic factors at the amide fragment of a new series of (+/-)-7a-alkyl-2-[4-(4-arylpiperazin-1-yl)butyl]-1,3-dioxoperhydropyrrolo[1,2-c]imidazoles 2 and of (+/-)-7a-alkyl-2-[(4-arylpiperazin-1-yl)methyl]-1,3-dioxoperhydropyrrolo[1,2-c]imidazoles 3 has been studied. Variations of logP have been carried out by introducing different hydrocarbonated substituents (R(1)) at the position 7a of the bicyclohydantoin, namely the non-pharmacophoric part. All the new compounds exhibit high potency for the 5-HT(1A) receptor; however, affinities for the alpha(1) receptor are high for compounds 2a-l while compounds 3a-f are selective over this adrenergic receptor. On the other hand, differences in logP do not notably affect the K(i) values for the above receptors. PMID:15018929

  19. Improved native affinity purification of RNA.

    PubMed

    Batey, Robert T; Kieft, Jeffrey S

    2007-08-01

    RNA biochemical or structural studies often require an RNA sample that is chemically pure, and most protocols for its in vitro production use denaturing polyacrylamide gel electrophoresis to achieve this. Unfortunately, many RNAs do not quantitatively refold into an active conformation after denaturation, creating significant problems for downstream characterization or use. In addition, this traditional purification method is not amenable to studies demanding high-throughput RNA production. Recently, we presented the first general method for producing almost any RNA sequence that employs an affinity tag that is removed during the purification process. Because technical difficulties prevented application of this method to many RNAs, we have developed an improved version that utilizes a different activatable ribozyme and affinity tag that are considerably more robust, rapid, and broadly applicable. PMID:17548432

  20. Protein affinity map of chemical space.

    PubMed

    Kauvar, L M; Villar, H O; Sportsman, J R; Higgins, D L; Schmidt, D E

    1998-09-11

    Affinity fingerprinting is a quantitative method for mapping chemical space based on binding preferences of compounds for a reference panel of proteins. An effective reference panel of <20 proteins can be empirically selected which shows differential interaction with nearly all compounds. By using this map to iteratively sample the chemical space, identification of active ligands from a library of 30,000 candidate compounds has been accomplished for a wide spectrum of specific protein targets. In each case, <200 compounds were directly assayed against the target. Further, analysis of the fingerprint database suggests a strategy for effective selection of affinity chromatography ligands and scaffolds for combinatorial chemistry. With such a system, the large numbers of potential therapeutic targets emerging from genome research can be categorized according to ligand binding properties, complementing sequence based classification. PMID:9792501

  1. Parameterization of an effective potential for protein–ligand binding from host–guest affinity data

    PubMed Central

    Wickstrom, Lauren; Deng, Nanjie; He, Peng; Mentes, Ahmet; Nguyen, Crystal; Gilson, Michael K.; Kurtzman, Tom; Gallicchio, Emilio; Levy, Ronald M.

    2015-01-01

    Force field accuracy is still one of the “stalemates” in biomolecular modeling. Model systems with high quality experimental data are valuable instruments for the validation and improvement of effective potentials. With respect to protein–ligand binding, organic host–guest complexes have long served as models for both experimental and computational studies because of the abundance of binding affinity data available for such systems. Binding affinity data collected for cyclodextrin (CD) inclusion complexes, a popular model for molecular recognition, is potentially a more reliable resource for tuning energy parameters than hydration free energy measurements. Convergence of binding free energy calculations on CD host–guest systems can also be obtained rapidly, thus offering the opportunity to assess the robustness of these parameters. In this work, we demonstrate how implicit solvent parameters can be developed using binding affinity experimental data and the binding energy distribution analysis method (BEDAM) and validated using the Grid Inhomogeneous Solvation Theory analysis. These new solvation parameters were used to study protein–ligand binding in two drug targets against the HIV-1 virus and improved the agreement between the calculated and the experimental binding affinities. This work illustrates how benchmark sets of high quality experimental binding affinity data and physics-based binding free energy models can be used to evaluate and optimize force fields for protein–ligand systems. PMID:26256816

  2. A MEMS Dielectric Affinity Glucose Biosensor.

    PubMed

    Huang, Xian; Li, Siqi; Davis, Erin; Li, Dachao; Wang, Qian; Lin, Qiao

    2013-06-20

    Continuous glucose monitoring (CGM) sensors based on affinity detection are desirable for long-term and stable glucose management. However, most affinity sensors contain mechanical moving structures and complex design in sensor actuation and signal readout, limiting their reliability in subcutaneously implantable glucose detection. We have previously demonstrated a proof-of-concept dielectric glucose sensor that measured pre-mixed glucose-sensitive polymer solutions at various glucose concentrations. This sensor features simplicity in sensor design, and possesses high specificity and accuracy in glucose detection. However, lack of glucose diffusion passage, this device is unable to fulfill real-time in-vivo monitoring. As a major improvement to this device, we present in this paper a fully implantable MEMS dielectric affinity glucose biosensor that contains a perforated electrode embedded in a suspended diaphragm. This capacitive-based sensor contains no moving parts, and enables glucose diffusion and real-time monitoring. The experimental results indicate that this sensor can detect glucose solutions at physiological concentrations and possesses good reversibility and reliability. This sensor has a time constant to glucose concentration change at approximately 3 min, which is comparable to commercial systems. The sensor has potential applications in fully implantable CGM that require excellent long-term stability and reliability. PMID:24511215

  3. On constructing purely affine theories with matter

    NASA Astrophysics Data System (ADS)

    Cervantes-Cota, Jorge L.; Liebscher, D.-E.

    2016-08-01

    We explore ways to obtain the very existence of a space-time metric from an action principle that does not refer to it a priori. Although there are reasons to believe that only a non-local theory can viably achieve this goal, we investigate here local theories that start with Schrödinger's purely affine theory (Schrödinger in Space-time structure. Cambridge UP, Cambridge, 1950), where he gave reasons to set the metric proportional to the Ricci curvature aposteriori. When we leave the context of unified field theory, and we couple the non-gravitational matter using some weak equivalence principle, we can show that the propagation of shock waves does not define a lightcone when the purely affine theory is local and avoids the explicit use of the Ricci tensor in realizing the weak equivalence principle. When the Ricci tensor is substituted for the metric, the equations seem to have only a very limited set of solutions. This backs the conviction that viable purely affine theories have to be non-local.

  4. Phosphopeptide Enrichment by Immobilized Metal Affinity Chromatography.

    PubMed

    Thingholm, Tine E; Larsen, Martin R

    2016-01-01

    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively charged metal ions such as Fe(3+), Ga(3+), Al(3+), Zr(4+), and Ti(4+) has made it possible to enrich phosphorylated peptides from peptide samples. However, the selectivity of most of the metal ions is limited, when working with highly complex samples, e.g., whole-cell extracts, resulting in contamination from nonspecific binding of non-phosphorylated peptides. This problem is mainly caused by highly acidic peptides that also share high binding affinity towards these metal ions. By lowering the pH of the loading buffer nonspecific binding can be reduced significantly, however with the risk of reducing specific binding capacity. After binding, the enriched phosphopeptides are released from the metal ions using alkaline buffers of pH 10-11, EDTA, or phosphate-containing buffers. Here we describe a protocol for IMAC using Fe(3+) for phosphopeptide enrichment. The principles are illustrated on a semi-complex peptide mixture. PMID:26584922

  5. Localization of Free Field Realizations of Affine Lie Algebras

    NASA Astrophysics Data System (ADS)

    Futorny, Vyacheslav; Grantcharov, Dimitar; Martins, Renato A.

    2015-04-01

    We use localization technique to construct new families of irreducible modules of affine Kac-Moody algebras. In particular, localization is applied to the first free field realization of the affine Lie algebra or, equivalently, to imaginary Verma modules.

  6. Improving Binding Affinity and Selectivity of Computationally Designed Ligand-Binding Proteins Using Experiments.

    PubMed

    Tinberg, Christine E; Khare, Sagar D

    2016-01-01

    The ability to de novo design proteins that can bind small molecules has wide implications for synthetic biology and medicine. Combining computational protein design with the high-throughput screening of mutagenic libraries of computationally designed proteins is emerging as a general approach for creating binding proteins with programmable binding modes, affinities, and selectivities. The computational step enables the creation of a binding site in a protein that otherwise does not (measurably) bind the intended ligand, and targeted mutagenic screening allows for validation and refinement of the computational model as well as provides orders-of-magnitude increases in the binding affinity. Deep sequencing of mutagenic libraries can provide insights into the mutagenic binding landscape and enable further affinity improvements. Moreover, in such a combined computational-experimental approach where the binding mode is preprogrammed and iteratively refined, selectivity can be achieved (and modulated) by the placement of specified amino acid side chain groups around the ligand in defined orientations. Here, we describe the experimental aspects of a combined computational-experimental approach for designing-using the software suite Rosetta-proteins that bind a small molecule of choice and engineering, using fluorescence-activated cell sorting and high-throughput yeast surface display, high affinity and ligand selectivity. We illustrated the utility of this approach by performing the design of a selective digoxigenin (DIG)-binding protein that, after affinity maturation, binds DIG with picomolar affinity and high selectivity over structurally related steroids. PMID:27094290

  7. Structure-based protocol for identifying mutations that enhance protein-protein binding affinities.

    PubMed

    Sammond, Deanne W; Eletr, Ziad M; Purbeck, Carrie; Kimple, Randall J; Siderovski, David P; Kuhlman, Brian

    2007-08-31

    The ability to manipulate protein binding affinities is important for the development of proteins as biosensors, industrial reagents, and therapeutics. We have developed a structure-based method to rationally predict single mutations at protein-protein interfaces that enhance binding affinities. The protocol is based on the premise that increasing buried hydrophobic surface area and/or reducing buried hydrophilic surface area will generally lead to enhanced affinity if large steric clashes are not introduced and buried polar groups are not left without a hydrogen bond partner. The procedure selects affinity enhancing point mutations at the protein-protein interface using three criteria: (1) the mutation must be from a polar amino acid to a non-polar amino acid or from a non-polar amino acid to a larger non-polar amino acid, (2) the free energy of binding as calculated with the Rosetta protein modeling program should be more favorable than the free energy of binding calculated for the wild-type complex and (3) the mutation should not be predicted to significantly destabilize the monomers. The performance of the computational protocol was experimentally tested on two separate protein complexes; Galpha(i1) from the heterotrimeric G-protein system bound to the RGS14 GoLoco motif, and the E2, UbcH7, bound to the E3, E6AP from the ubiquitin pathway. Twelve single-site mutations that were predicted to be stabilizing were synthesized and characterized in the laboratory. Nine of the 12 mutations successfully increased binding affinity with five of these increasing binding by over 1.0 kcal/mol. To further assess our approach we searched the literature for point mutations that pass our criteria and have experimentally determined binding affinities. Of the eight mutations identified, five were accurately predicted to increase binding affinity, further validating the method as a useful tool to increase protein-protein binding affinities. PMID:17603074

  8. A strategy of designing the ligand of antibody affinity chromatography based on molecular dynamics simulation.

    PubMed

    Dai, Lu; Li, Weikang; Sun, Fei; Li, Baizhi; Li, Hongrui; Zhang, Hongxing; Zheng, Qingchuan; Liang, Chongyang

    2016-09-01

    Designing affinity ligands has always been the development focus of affinity chromatography. Previous antibody affinity ligand designs were mostly based on the crystal structure of protein A (UniProt code number: P38507), and the antibody-binding domains were modified according to the properties of amino acid residues. Currently, more effective bioinformatic prediction and experimental validation has been used to improve the design of antibody affinity ligands. In the present study, the complex crystal structure (the domain D of protein A and the Fab segment of IgM, PDB code: 1DEE) was used as the model. The vital site that inhibits the binding between domain D and IgM was estimated by means of molecular dynamics (MD) simulation, then MM-GBSA calculations were used to design a mutant of domain D (K46E) for improving affinity on the above vital site. The binding analysis using Biacore showed the association and dissociation parameters of K46E mutant that were optimized with IgM. The affinity increase of K46E mutant preferred for IgM, the affinity order is K46E tetramer (KD=6.02×10(-9)M)>K46E mutant (KD=6.66×10(-8)M)>domain D (KD=2.17×10(-7)M). Similar results were obtained when the optimized ligands were immobilized to the chromatography medium. A complete designing strategy was validated in this study, which will provide a novel insight into designing new ligands of antibody affinity chromatography media. PMID:27524303

  9. The affinity of magnetic microspheres for Schistosoma eggs.

    PubMed

    Candido, Renata R F; Favero, Vivian; Duke, Mary; Karl, Stephan; Gutiérrez, Lucía; Woodward, Robert C; Graeff-Teixeira, Carlos; Jones, Malcolm K; St Pierre, Timothy G

    2015-01-01

    Schistosomiasis is a chronic parasitic disease of humans, with two species primarily causing the intestinal infection: Schistosoma mansoni and Schistosoma japonicum. Traditionally, diagnosis of schistosomiasis is achieved through direct visualisation of eggs in faeces using techniques that lack the sensitivity required to detect all infections, especially in areas of low endemicity. A recently developed method termed Helmintex™ is a very sensitive technique for detection of Schistosoma eggs and exhibits 100% sensitivity at 1.3 eggs per gram of faeces, enough to detect even low-level infections. The Helminthex™ method is based on the interaction of magnetic microspheres and schistosome eggs. Further understanding the underlying egg-microsphere interactions would enable a targeted optimisation of egg-particle binding and may thus enable a significant improvement of the Helmintex™ method and diagnostic sensitivity in areas with low infection rates. We investigated the magnetic properties of S. mansoni and S. japonicum eggs and their interactions with microspheres with different magnetic properties and surface functionalization. Eggs of both species exhibited higher binding affinity to the magnetic microspheres than the non-magnetic microspheres. Binding efficiency was further enhanced if the particles were coated with streptavidin. Schistosoma japonicum eggs bound more microspheres compared with S. mansoni. However, distinct differences within eggs of each species were also observed when the distribution of the number of microspheres bound per egg was modelled with double Poisson distributions. Using this approach, both S. japonicum and S. mansoni eggs fell into two groups, one having greater affinity for magnetic microspheres than the other, indicating that not all eggs of a species exhibit the same binding affinity. Our observations suggest that interaction between the microspheres and eggs is more likely to be related to surface charge-based electrostatic

  10. Dicyanovinylnaphthalenes for neuroimaging of amyloids and relationships of electronic structures and geometries to binding affinities

    PubMed Central

    Petrič, Andrej; Johnson, Scott A.; Pham, Hung V.; Li, Ying; Čeh, Simon; Golobič, Amalija; Agdeppa, Eric D.; Timbol, Gerald; Liu, Jie; Keum, Gyochang; Satyamurthy, Nagichettiar; Kepe, Vladimir; Houk, Kendall N.; Barrio, Jorge R.

    2012-01-01

    The positron-emission tomography (PET) probe 2-(1-[6-[(2-fluoroethyl)(methyl)amino]-2-naphthyl]ethylidene) (FDDNP) is used for the noninvasive brain imaging of amyloid-β (Aβ) and other amyloid aggregates present in Alzheimer’s disease and other neurodegenerative diseases. A series of FDDNP analogs has been synthesized and characterized using spectroscopic and computational methods. The binding affinities of these molecules have been measured experimentally and explained through the use of a computational model. The analogs were created by systematically modifying the donor and the acceptor sides of FDDNP to learn the structural requirements for optimal binding to Aβ aggregates. FDDNP and its analogs are neutral, environmentally sensitive, fluorescent molecules with high dipole moments, as evidenced by their spectroscopic properties and dipole moment calculations. The preferred solution-state conformation of these compounds is directly related to the binding affinities. The extreme cases were a nonplanar analog t-butyl-FDDNP, which shows low binding affinity for Aβ aggregates (520 nM Ki) in vitro and a nearly planar tricyclic analog cDDNP, which displayed the highest binding affinity (10 pM Ki). Using a previously published X-ray crystallographic model of 1,1-dicyano-2-[6-(dimethylamino)naphthalen-2-yl]propene (DDNP) bound to an amyloidogenic Aβ peptide model, we show that the binding affinity is inversely related to the distortion energy necessary to avoid steric clashes along the internal surface of the binding channel. PMID:23012452

  11. Measuring an antibody affinity distribution molecule by molecule

    SciTech Connect

    Bradbury, Andrew M; Werner, James H; Temirov, Jamshid

    2008-01-01

    Single molecule fluorescence mIcroscopy was used to observe the binding and unbinding of hapten decorated quantum dots with individual surface immobilized antibodies. The fluorescence time history from an individual antibody site can be used to calculate its binding affinity. While quantum dot blinking occurs during these measurements, we describe a simple empirical method to correct the apparent/observed affinity to account for the blinking contribution. The combination of many single molecule affinity measurements from different antibodies yields not only the average affinity, it directly measures the full shape and character of the surface affinity distribution function.

  12. On the structure of self-affine convex bodies

    SciTech Connect

    Voynov, A S

    2013-08-31

    We study the structure of convex bodies in R{sup d} that can be represented as a union of their affine images with no common interior points. Such bodies are called self-affine. Vallet's conjecture on the structure of self-affine bodies was proved for d = 2 by Richter in 2011. In the present paper we disprove the conjecture for all d≥3 and derive a detailed description of self-affine bodies in R{sup 3}. Also we consider the relation between properties of self-affine bodies and functional equations with a contraction of an argument. Bibliography: 10 titles.

  13. Metal-affinity separations: A new dimension in protein processing

    SciTech Connect

    Arnold, F.H. )

    1991-02-01

    Rapid growth in the preparative and high-resolution analytical applications of metal-affinity chromatography demonstrate the appeal of metal recognition as a basis for protein separations. Stable, inexpensive chelated metals effectively mimic biospecific interactions, providing selective ligands for protein binding. This article reviews recent progress in understanding the mechanisms of metal-protein recognition that underlie metal-affinity separations. Also discussed are schemes for integrating metal-affinity purifications into the expression and bioprocessing of recombinant proteins. Promising future developments include new metal-affinity processes for analytical and preparative-scale separations and a range of techniques for enhancing the selectivity of metal-affinity separations.

  14. Avoiding degenerate coframes in an affine gauge approach to quantum gravity

    SciTech Connect

    Mielke, E.W.; McCrea, J.D.; Ne`eman, Y.; Hehl, F.W.

    1993-04-01

    This report discusses the following concepts on quantum gravity: The affine gauge approach; affine gauge transformations versus active differomorphisms; affine gauge approach to quantum gravity with topology change.

  15. Aluminum monocation basicity and affinity scales.

    PubMed

    Gal, Jean-François; Yáñez, Manuel; Mó, Otilia

    2015-01-01

    The experimental aspects of the determination of thermochemical data for the attachment of the aluminum monocation Al(+) to neutral atoms and molecules are reviewed. Literature aluminum cation affinities (enthalpy scale) and basicities (Gibbs energy scale) are tabulated and discussed. Ab initio quantum chemical calculations at the G4 level on 43 adducts provide a consistent picture of the energetics of the adducts and their structures. The Al(+)-ligand bonding is analyzed in terms of natural bond orbital and atom-in molecule analyses. A brief comparison of the Al(+) basicity scales and other gas- phase cation basicities is presented. PMID:26307732

  16. Contractions of affine Kac-Moody algebras

    NASA Astrophysics Data System (ADS)

    Daboul, J.; Daboul, C.; de Montigny, M.

    2008-08-01

    I review our recent work on contractions of affine Kac-Moody algebras (KMA) and present new results. We study generalized contractions of KMA with respect to their twisted and untwisted KM subalgebras. As a concrete example, we discuss contraction of D(1)4 and D(3)4, based on Z3-grading. We also describe examples of 'level-dependent' contractions, which are based on Z-gradings of KMA. Our work generalizes the Inönü-Wigner contraction of P. Majumdar in several directions. We also give an algorithm for constructing Kac-Moody-like algebras hat g for any Lie algebra g.

  17. Aptamer Affinity Maturation by Resampling and Microarray Selection.

    PubMed

    Kinghorn, Andrew B; Dirkzwager, Roderick M; Liang, Shaolin; Cheung, Yee-Wai; Fraser, Lewis A; Shiu, Simon Chi-Chin; Tang, Marco S L; Tanner, Julian A

    2016-07-19

    Aptamers have significant potential as affinity reagents, but better approaches are critically needed to discover higher affinity nucleic acids to widen the scope for their diagnostic, therapeutic, and proteomic application. Here, we report aptamer affinity maturation, a novel aptamer enhancement technique, which combines bioinformatic resampling of aptamer sequence data and microarray selection to navigate the combinatorial chemistry binding landscape. Aptamer affinity maturation is shown to improve aptamer affinity by an order of magnitude in a single round. The novel aptamers exhibited significant adaptation, the complexity of which precludes discovery by other microarray based methods. Honing aptamer sequences using aptamer affinity maturation could help optimize a next generation of nucleic acid affinity reagents. PMID:27346322

  18. Probing a potential in vivo drug-excipient interaction: temporal effects of a modified β-cyclodextrin on the intravenous pharmacokinetics of a model high-affinity drug ligand.

    PubMed

    Mannila, Anne; Morizzi, Julia; Nguyen, Thuy Tien; Charman, Susan A; McIntosh, Michelle P; Shackleford, David M

    2012-09-01

    The investigational synthetic ozonide, OZ209, has previously been shown to have high binding affinity for sulfobutylether(7)-β-cyclodextrin [(SBE)(7)-β-CD] resulting in altered pharmacokinetics when administered intravenously to rats in a (SBE)(7)-β-CD aqueous formulation. In the present study, OZ209 and (SBE)(7)-β-CD have been used to probe whether a modified β-CD excipient, on systemic administration, can bind to and alter the pharmacokinetics of a coadministered drug. When (SBE)(7)-β-CD was administered 60 min after OZ209, a spike in the concentration of OZ209 in blood and plasma was detected within 2 min of the (SBE)(7)-β-CD infusion, and this was accompanied by a temporary decrease in the whole blood-to-plasma partitioning ratio of OZ209, the duration of which was dependent upon the dose of (SBE)(7)-β-CD. Administration of (SBE)(7)-β-CD also resulted in increased urinary excretion of OZ209. By contrast, administration of (SBE)(7)-β-CD 4 h prior to OZ209 had no pronounced effect on the blood or plasma pharmacokinetics of OZ209, consistent with the (SBE)(7)-β-CD having been largely eliminated prior to the administration of OZ209. This study is the first to demonstrate an in vivo drug-excipient interaction between a modified β-CD and a coadministered drug, and also demonstrates that such an interaction can be avoided through appropriate consideration of CD pharmacokinetics. PMID:22549698

  19. 01-ERD-111 - The Development of Synthetic High Affinity Ligands

    SciTech Connect

    Perkins, J; Balhorn, R; Cosman, M; Lightstone, F; Zeller, L

    2004-02-05

    The aim of this project was to develop Synthetic High-Affinity Ligands (SHALs), which bind with high affinity and specificity to proteins of interest for national security and cancer therapy applications. The aim of producing synthetic ligands for sensory devices as an alternative to antibody-based detection assays and therapeutic agents is to overcome the drawbacks associated with antibody-based in next-generation sensors and systems. The focus area of the project was the chemical synthesis of the SHALs. The project concentrated on two different protein targets. (a) The C fragment of tetanus and botulinum toxin, potential biowarfare agents. A SHAL for tetanus or botulinum toxin would be incorporated into a sensory device for the toxins. (b) HLA-DR10, a protein found in high abundance on the surface of Non-Hodgkins Lymphoma. A SHAL specific to a tumor marker, labeled with a radionuclide, would enable the targeted delivery of radiation therapy to metastatic disease. The technical approach used to develop a SHAL for each protein target will be described in more detail below. However, in general, the development of a SHAL requires a combination of computational modeling techniques, modern nuclear magnetic resonance spectroscopy (NMR) and synthetic chemistry.

  20. Expression and affinity purification of recombinant proteins from plants

    NASA Technical Reports Server (NTRS)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  1. High-affinity Cyclic Peptide Matriptase Inhibitors*

    PubMed Central

    Quimbar, Pedro; Malik, Uru; Sommerhoff, Christian P.; Kaas, Quentin; Chan, Lai Y.; Huang, Yen-Hua; Grundhuber, Maresa; Dunse, Kerry; Craik, David J.; Anderson, Marilyn A.; Daly, Norelle L.

    2013-01-01

    The type II transmembrane serine protease matriptase is a key activator of multiple signaling pathways associated with cell proliferation and modification of the extracellular matrix. Deregulated matriptase activity correlates with a number of diseases, including cancer and hence highly selective matriptase inhibitors may have therapeutic potential. The plant-derived cyclic peptide, sunflower trypsin inhibitor-1 (SFTI-1), is a promising drug scaffold with potent matriptase inhibitory activity. In the current study we have analyzed the structure-activity relationships of SFTI-1 and Momordica cochinchinensis trypsin inhibitor-II (MCoTI-II), a structurally divergent trypsin inhibitor from Momordica cochinchinensis that also contains a cyclic backbone. We show that MCoTI-II is a significantly more potent matriptase inhibitor than SFTI-1 and that all alanine mutants of both peptides, generated using positional scanning mutagenesis, have decreased trypsin affinity, whereas several mutations either maintain or result in enhanced matriptase inhibitory activity. These intriguing results were used to design one of the most potent matriptase inhibitors known to date with a 290 pm equilibrium dissociation constant, and provide the first indication on how to modulate affinity for matriptase over trypsin in cyclic peptides. This information might be useful for the design of more selective and therapeutically relevant inhibitors of matriptase. PMID:23548907

  2. Heparin affinity purification of extracellular vesicles

    PubMed Central

    Balaj, Leonora; Atai, Nadia A.; Chen, Weilin; Mu, Dakai; Tannous, Bakhos A.; Breakefield, Xandra O.; Skog, Johan; Maguire, Casey A.

    2015-01-01

    Extracellular vesicles (EVs) are lipid membrane vesicles released by cells. They carry active biomolecules including DNA, RNA, and protein which can be transferred to recipient cells. Isolation and purification of EVs from culture cell media and biofluids is still a major challenge. The most widely used isolation method is ultracentrifugation (UC) which requires expensive equipment and only partially purifies EVs. Previously we have shown that heparin blocks EV uptake in cells, supporting a direct EV-heparin interaction. Here we show that EVs can be purified from cell culture media and human plasma using ultrafiltration (UF) followed by heparin-affinity beads. UF/heparin-purified EVs from cell culture displayed the EV marker Alix, contained a diverse RNA profile, had lower levels of protein contamination, and were functional at binding to and uptake into cells. RNA yield was similar for EVs isolated by UC. We were able to detect mRNAs in plasma samples with comparable levels to UC samples. In conclusion, we have discovered a simple, scalable, and effective method to purify EVs taking advantage of their heparin affinity. PMID:25988257

  3. Affine conformal vectors in space-time

    NASA Astrophysics Data System (ADS)

    Coley, A. A.; Tupper, B. O. J.

    1992-05-01

    All space-times admitting a proper affine conformal vector (ACV) are found. By using a theorem of Hall and da Costa, it is shown that such space-times either (i) admit a covariantly constant vector (timelike, spacelike, or null) and the ACV is the sum of a proper affine vector and a conformal Killing vector or (ii) the space-time is 2+2 decomposable, in which case it is shown that no ACV can exist (unless the space-time decomposes further). Furthermore, it is proved that all space-times admitting an ACV and a null covariantly constant vector (which are necessarily generalized pp-wave space-times) must have Ricci tensor of Segré type {2,(1,1)}. It follows that, among space-times admitting proper ACV, the Einstein static universe is the only perfect fluid space-time, there are no non-null Einstein-Maxwell space-times, and only the pp-wave space-times are representative of null Einstein-Maxwell solutions. Otherwise, the space-times can represent anisotropic fluids and viscous heat-conducting fluids, but only with restricted equations of state in each case.

  4. Exploring Fluorous Affinity by Liquid Chromatography.

    PubMed

    Catani, Martina; Guzzinati, Roberta; Marchetti, Nicola; Pasti, Luisa; Cavazzini, Alberto

    2015-07-01

    Terms such as "fluorous affinity" and "fluorophilicity" have been used to describe the unique partition and sorption properties often exhibited by highly fluorinated organic compounds, that is molecules rich in sp(3) carbon-fluorine bonds. In this work, we made use of a highly fluorinated stationary phase and a series of benzene derivatives to study the effect of one single perfluorinated carbon on the chromatographic behavior and adsorption properties of molecules. For this purpose, the adsorption equilibria of α,α,α-trifluorotoluene, toluene, and other alkylbenzenes have been studied by means of nonlinear chromatography in a variety of acetonitrile/water eluents. Our results reveal that one single perfluorinated carbon is already enough to induce a drastic change in the adsorption properties of molecules on the perfluorinated stationary phase. In particular, it has been found that adsorption is monolayer if the perfluoroalkyl carbon is present but that, when this unit is missing, molecules arrange as multilayer stack structures. These findings can contribute to the understanding of molecular mechanisms of fluorous affinity. PMID:26047527

  5. Quantification of hydrophobic interaction affinity of colloids

    NASA Astrophysics Data System (ADS)

    Saini, G.; Nasholm, N.; Wood, B. D.

    2009-12-01

    Colloids play an important role in a wide variety of disciplines, including water and wastewater treatment, subsurface transport of metals and organic contaminants, migration of fines in oil reservoirs, biocolloid (virus and bacteria) transport in subsurface, and are integral to laboratory transport studies. Although the role of hydrophobicity in adhesion and transport of colloids, particularly bacteria, is well known; there is scarcity of literature regarding hydrophobicity measurement of non-bacterial colloids and other micron-sized particles. Here we detail an experimental approach based on differential partitioning of colloids between two liquid phases (hydrocarbon and buffer) as a measure of the hydrophobic interaction affinity of colloids. This assay, known as Microbial adhesion to hydrocarbons or MATH, is frequently used in microbiology and bacteriology for quantifying the hydrophobicity of microbes. Monodispersed colloids and particles, with sizes ranging from 1 micron to 33 micron, were used for the experiments. A range of hydrophobicity values were observed for different particles. The hydrophobicity results are also verified against water contact angle measurements of these particles. This liquid-liquid partitioning assay is quick, easy-to-perform and requires minimal instrumentation. Estimation of the hydrophobic interaction affinity of colloids would lead to a better understanding of their adhesion to different surfaces and subsequent transport in porous media.

  6. Impedance-derived electrochemical capacitance spectroscopy for the evaluation of lectin-glycoprotein binding affinity.

    PubMed

    Santos, Adriano; Carvalho, Fernanda C; Roque-Barreira, Maria-Cristina; Bueno, Paulo R

    2014-12-15

    Characterization of lectin-carbohydrate binding using label-free methods such as impedance-derived electrochemical capacitance spectroscopy (ECS) is desirable to evaluate specific interactions, for example, ArtinM lectin and horseradish peroxidase (HRP) glycoprotein, used here as a model for protein-carbohydrate binding affinity. An electroactive molecular film comprising alkyl ferrocene as a redox probe and ArtinM as a carbohydrate receptive center to target HRP was successfully used to determine the binding affinity between ArtinM and HRP. The redox capacitance, a transducer signal associated with the alkyl ferrocene centers, was obtained by ECS and used in the Langmuir adsorption model to obtain the affinity constant (1.6±0.6)×10(8) L mol(-1). The results shown herein suggest the feasibility of ECS application for lectin glycoarray characterization. PMID:24994505

  7. Soybean. beta. -glucan binding sites display maximal affinity for a heptaglucoside phytoalexin-elicitor

    SciTech Connect

    Cosio, E.G.; Waldmueller, T.; Frey, T.; Ebel, J. )

    1990-05-01

    The affinity of soybean {beta}-glucan-binding sites for a synthetic heptaglucan elicitor was tested in a ligand-competition assay against a {sup 125}I-labeled 1,3-1,6-{beta}-glucan preparation (avg. DP=20). Half-maximal displacement of label (IC{sub 50}) was obtained at 9nM heptaglucan, the highest affinity of all fractions tested to date. Displacement followed a uniform sigmoidal pattern and was complete at 1{mu}M indicating access of heptaglucan to all sites available to the labeled elicitor. A mathematical model was used to predict IC{sub 50} values according to the DP of glucan fragments obtained from fungal cell walls. The lowest IC{sub 50} predicted by this model is 3nM. Binding affinity of the glucans was compared with their elicitor activity in a bioassay.

  8. On the binding affinity of macromolecular interactions: daring to ask why proteins interact

    PubMed Central

    Kastritis, Panagiotis L.; Bonvin, Alexandre M. J. J.

    2013-01-01

    Interactions between proteins are orchestrated in a precise and time-dependent manner, underlying cellular function. The binding affinity, defined as the strength of these interactions, is translated into physico-chemical terms in the dissociation constant (Kd), the latter being an experimental measure that determines whether an interaction will be formed in solution or not. Predicting binding affinity from structural models has been a matter of active research for more than 40 years because of its fundamental role in drug development. However, all available approaches are incapable of predicting the binding affinity of protein–protein complexes from coordinates alone. Here, we examine both theoretical and experimental limitations that complicate the derivation of structure–affinity relationships. Most work so far has concentrated on binary interactions. Systems of increased complexity are far from being understood. The main physico-chemical measure that relates to binding affinity is the buried surface area, but it does not hold for flexible complexes. For the latter, there must be a significant entropic contribution that will have to be approximated in the future. We foresee that any theoretical modelling of these interactions will have to follow an integrative approach considering the biology, chemistry and physics that underlie protein–protein recognition. PMID:23235262

  9. Affinity Crystallography: A New Approach to Extracting High-Affinity Enzyme Inhibitors from Natural Extracts.

    PubMed

    Aguda, Adeleke H; Lavallee, Vincent; Cheng, Ping; Bott, Tina M; Meimetis, Labros G; Law, Simon; Nguyen, Nham T; Williams, David E; Kaleta, Jadwiga; Villanueva, Ivan; Davies, Julian; Andersen, Raymond J; Brayer, Gary D; Brömme, Dieter

    2016-08-26

    Natural products are an important source of novel drug scaffolds. The highly variable and unpredictable timelines associated with isolating novel compounds and elucidating their structures have led to the demise of exploring natural product extract libraries in drug discovery programs. Here we introduce affinity crystallography as a new methodology that significantly shortens the time of the hit to active structure cycle in bioactive natural product discovery research. This affinity crystallography approach is illustrated by using semipure fractions of an actinomycetes culture extract to isolate and identify a cathepsin K inhibitor and to compare the outcome with the traditional assay-guided purification/structural analysis approach. The traditional approach resulted in the identification of the known inhibitor antipain (1) and its new but lower potency dehydration product 2, while the affinity crystallography approach led to the identification of a new high-affinity inhibitor named lichostatinal (3). The structure and potency of lichostatinal (3) was verified by total synthesis and kinetic characterization. To the best of our knowledge, this is the first example of isolating and characterizing a potent enzyme inhibitor from a partially purified crude natural product extract using a protein crystallographic approach. PMID:27498895

  10. Structure of a High-Affinity

    SciTech Connect

    Saphire, E.O.; Montero, M.; Menendez, A.; Houten, N.E.van; Irving, M.B.; Pantophlet, R.; Swick, M.B.; Parren, P.W.H.I.; Burton, D.R.; Scott, J.K.; Wilson, I.A.; /Scripps Res. Inst. /Simon Fraser U. /British Columbia U.

    2007-07-13

    The human antibody b12 recognizes a discontinuous epitope on gp120 and is one of the rare monoclonal antibodies that neutralize a broad range of primary human immunodeficiency virus type 1 (HIV-1) isolates. We previously reported the isolation of B2.1, a dimeric peptide that binds with high specificity to b12 and competes with gp120 for b12 antibody binding. Here, we show that the affinity of B2.1 was improved 60-fold over its synthetic-peptide counterpart by fusing it to the N terminus of a soluble protein. This affinity, which is within an order of magnitude of that of gp120, probably more closely reflects the affinity of the phage-borne peptide. The crystal structure of a complex between Fab of b12 and B2.1 was determined at 1.8 Angstrom resolution. The structural data allowed the differentiation of residues that form critical contacts with b12 from those required for maintenance of the antigenic structure of the peptide, and revealed that three contiguous residues mediate B2.1's critical contacts with b12. This single region of critical contact between the B2.1 peptide and the b12 paratope is unlikely to mimic the discontinuous key binding residues involved in the full b12 epitope for gp120, as previously identified by alanine scanning substitutions on the gp120 surface. These structural observations are supported by experiments that demonstrate that B2.1 is an ineffective immunogenic mimic of the b12 epitope on gp120. Indeed, an extensive series of immunizations with B2.1 in various forms failed to produce gp120 cross-reactive sera. The functional and structural data presented here, however, suggest that the mechanism by which b12 recognizes the two antigens is very different. Here, we present the first crystal structure of peptide bound to an antibody that was originally raised against a discontinuous protein epitope. Our results highlight the challenge of producing immunogens that mimic discontinuous protein epitopes, and the necessity of combining

  11. Automatic gesture analysis using constant affine velocity.

    PubMed

    Cifuentes, Jenny; Boulanger, Pierre; Pham, Minh Tu; Moreau, Richard; Prieto, Flavio

    2014-01-01

    Hand human gesture recognition has been an important research topic widely studied around the world, as this field offers the ability to identify, recognize, and analyze human gestures in order to control devices or to interact with computer interfaces. In particular, in medical training, this approach is an important tool that can be used to obtain an objective evaluation of a procedure performance. In this paper, some obstetrical gestures, acquired by a forceps, were studied with the hypothesis that, as the scribbling and drawing movements, they obey the one-sixth power law, an empirical relationship which connects path curvature, torsion, and euclidean velocity. Our results show that obstetrical gestures have a constant affine velocity, which is different for each type of gesture and based on this idea this quantity is proposed as an appropriate classification feature in the hand human gesture recognition field. PMID:25570332

  12. Effectively nonlocal metric-affine gravity

    NASA Astrophysics Data System (ADS)

    Golovnev, Alexey; Koivisto, Tomi; Sandstad, Marit

    2016-03-01

    In metric-affine theories of gravity such as the C-theories, the spacetime connection is associated to a metric that is nontrivially related to the physical metric. In this article, such theories are rewritten in terms of a single metric, and it is shown that they can be recast as effectively nonlocal gravity. With some assumptions, known ghost-free theories with nonsingular and cosmologically interesting properties may be recovered. Relations between different formulations are analyzed at both perturbative and nonperturbative levels, taking carefully into account subtleties with boundary conditions in the presence of integral operators in the action, and equivalences between theories related by nonlocal redefinitions of the fields are verified at the level of equations of motion. This suggests a possible geometrical interpretation of nonlocal gravity as an emergent property of non-Riemannian spacetime structure.

  13. Affinities of the Swartkrans early Homo mandibles.

    PubMed

    Curnoe, Darren

    2008-01-01

    The southern African early Homo assemblage continues to make important contributions to understanding the systematics, adaptations and evolutionary history of the human genus. However, the taxonomy of this sample is in a state of flux. This study examines the size and shape of the mandibular bodies of Swartkrans SK 15 and SK 45 comparing them with variation in two early Homo taxa (H. habilis sensu lato and H. sapiens erectus). The research aims to clarify their phenetic affinities and systematics through univariate statistics, inferential testing and multivariate analysis employing size (Log-transformed) and shape (Mosimann variables). Neither of them strongly resembles H. habilis sensu lato or H. sapiens erectus, rather, they probably sample a novel species of Homo not seen in East Africa. Moreover, there is considerable morphological variability within the Swartkrans sample and the possibility of more than one novel species being sampled at this site cannot be excluded. PMID:18402959

  14. Dye affinity cryogels for plasmid DNA purification.

    PubMed

    Çimen, Duygu; Yılmaz, Fatma; Perçin, Işık; Türkmen, Deniz; Denizli, Adil

    2015-11-01

    The aim of this study is to prepare megaporous dye-affinity cryogel discs for the purification of plasmid DNA (pDNA) from bacterial lysate. Poly(hydroxyethyl methacrylate) [PHEMA] cryogel discs were produced by free radical polymerization initiated by N,N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) redox pair in an ice bath. Cibacron Blue F3GA was used as an affinity ligand (loading amount: 68.9μmol/g polymer). The amount of pDNA adsorbed onto the PHEMA-Cibacron Blue F3GA cryogel discs first increased and then reached a plateau value (i.e., 32.5mg/g cryogel) at 3.0mg/mL pDNA concentration. Compared with the PHEMA cryogel (0.11mg/g cryogel), the pDNA adsorption capacity of the PHEMA-Cibacron Blue F3GA cryogel (32.4mg/g polymer) was improved significantly due to the Cibacron Blue 3GA immobilization onto the polymeric matrix. pDNA adsorption amount decreased from 11.7mg/g to 1.1mg/g with the increasing of NaCl concentration. The maximum pDNA adsorption was achieved at 4°C. The overall recovery of pDNA was calculated as 90%. The PHEMA-Cibacron Blue F3GA cryogel discs could be used five times without decreasing the pDNA adsorption capacity significantly. The results show that the PHEMA-Cibacron Blue F3GA cryogel discs promise high selectivity for pDNA. PMID:26249596

  15. In vitro affinity maturation of a natural human antibody overcomes a barrier to in vivo affinity maturation

    PubMed Central

    Li, Bing; Fouts, Ashley E; Stengel, Katharina; Luan, Peng; Dillon, Michael; Liang, Wei-Ching; Feierbach, Becket; Kelley, Robert F; Hötzel, Isidro

    2014-01-01

    Antibodies isolated from human donors are increasingly being developed for anti-infective therapeutics. These antibodies undergo affinity maturation in vivo, minimizing the need for engineering of therapeutic leads for affinity. However, the affinities required for some therapeutic applications may be higher than the affinities of the leads obtained, requiring further affinity maturation in vitro. To improve the neutralization potency of natural human antibody MSL-109 targeting human cytomegalovirus (CMV), we affinity matured the antibody against the gH/gL glycoprotein complex. A phage display library where most of the six complementary-determining regions (CDRs) were allowed to vary in only one amino acid residue at a time was used to scan for mutations that improve binding affinity. A T55R mutation and multiple mutations in position 53 of the heavy chain were identified that, when present individually or in combination, resulted in higher apparent affinities to gH/gL and improved CMV neutralization potency of Fab fragments expressed in bacterial cells. Three of these mutations in position 53 introduced glycosylation sites in heavy chain CDR 2 (CDR H2) that impaired binding of antibodies expressed in mammalian cells. One high affinity (KD < 10 pM) variant was identified that combined the D53N and T55R mutations while avoiding glycosylation of CDR H2. However, all the amino acid substitutions identified by phage display that improved binding affinity without introducing glycosylation sites required between two and four simultaneous nucleotide mutations to avoid glycosylation. These results indicate that the natural human antibody MSL-109 is close to a local affinity optimum. We show that affinity maturation by phage display can be used to identify and bypass barriers to in vivo affinity maturation of antibodies imposed by glycosylation and codon usage. These constraints may be relatively prevalent in human antibodies due to the codon usage and the amino acid

  16. Prediction of Neutral Salt Elution Profiles for Affinity Chromatography

    NASA Astrophysics Data System (ADS)

    Robinson, Jack B.; Strottmann, James M.; Stellwagen, Earle

    1981-04-01

    Neutral salts exhibit very marked differences as eluants of proteins from affinity columns. We observe: (i) that the relative potencies of neutral salts as eluants are independent of the protein or the affinity ligand in the systems studied, (ii) that the absolute salt concentration necessary to elute any given protein bound to the affinity matrix is proportional to the algebraic sum of a set of elution coefficients defined herein for the separate ions present in the solution, and (iii) that the proportionality between elution potency and elution coefficient is a function of the affinity of the protein for the immobilized ligand. Given the concentration of one neutral salt required for elution of a protein of interest from an affinity column, the elution capability of any neutral salt at any temperature can be quantitatively predicted for that protein. Accordingly, application and elution protocols for affinity chromatography can be designed to optimize the yield and fold purification of proteins.

  17. The derivatization of oxidized polysaccharides for protein immobilization and affinity chromatography.

    PubMed

    Junowicz, E; Charm, S E

    1976-03-25

    The present report describes the preparation of modified polysaccharides matrices useful for the synthesis of affinity adsorbents and immobilized proteins. Hydrazido-matrices were synthesized by condensing an excess of the bifunctional reagent, adipic acid dihydrazide, with periodate oxidized cellulose paper, Sephadex, or Sepharose matrices. Ribonucleotide dialdehyde cofactors, glyceraldehyde 3-phosphate, pyridoxal 5'-phosphate and oxidized DNAase B were separately bound to the hydrazido-polymers. Azido-matrices obtained by modification of the hydrazido-derivatives were coupled to specific amino ligands such as amino acids and proteins. Several adsorbents were prepared and used as models for affinity chromatography. PMID:1260016

  18. Vibrational photodetachment spectroscopy near the electron affinity of S2

    NASA Astrophysics Data System (ADS)

    Barrick, J. B.; Yukich, J. N.

    2016-02-01

    We have conducted laser photodetachment spectroscopy near the detachment threshold of the electron affinity of S2 in a 1.8-T field. The ions are prepared by dissociative electron attachment to carbonyl sulfide. The experiment is conducted in a Penning ion trap and with a narrow-band, tunable, Ti:sapphire laser. A hybrid model for photodetachment in an ion trap is fit to the data using the appropriate Franck-Condon factors. The observations reveal detachment from and to the first few vibrational levels of the anion and the neutral molecule, respectively. Evaporative cooling of the anion ensemble condenses the thermal distribution to the lowest initial vibrational states. The subsequent detachment spectroscopy yields results consistent with a vibrationally cooled anion population.

  19. 3D affine registration using teaching-learning based optimization

    NASA Astrophysics Data System (ADS)

    Jani, Ashish; Savsani, Vimal; Pandya, Abhijit

    2013-09-01

    3D image registration is an emerging research field in the study of computer vision. In this paper, two effective global optimization methods are considered for the 3D registration of point clouds. Experiments were conducted by applying each algorithm and their performance was evaluated with respect to rigidity, similarity and affine transformations. Comparison of algorithms and its effectiveness was tested for the average performance to find the global solution for minimizing the error in the terms of distance between the model cloud and the data cloud. The parameters for the transformation matrix were considered as the design variables. Further comparisons of the considered methods were done for the computational effort, computational time and the convergence of the algorithm. The results reveal that the use of TLBO was outstanding for image processing application involving 3D registration. [Figure not available: see fulltext.

  20. Affinity sensor based on immobilized molecular imprinted synthetic recognition elements.

    PubMed

    Lenain, Pieterjan; De Saeger, Sarah; Mattiasson, Bo; Hedström, Martin

    2015-07-15

    An affinity sensor based on capacitive transduction was developed to detect a model compound, metergoline, in a continuous flow system. This system simulates the monitoring of low-molecular weight organic compounds in natural flowing waters, i.e. rivers and streams. During operation in such scenarios, control of the experimental parameters is not possible, which poses a true analytical challenge. A two-step approach was used to produce a sensor for metergoline. Submicron spherical molecularly imprinted polymers, used as recognition elements, were obtained through emulsion polymerization and subsequently coupled to the sensor surface by electropolymerization. This way, a robust and reusable sensor was obtained that regenerated spontaneously under the natural conditions in a river. Small organic compounds could be analyzed in water without manipulating the binding or regeneration conditions, thereby offering a viable tool for on-site application. PMID:25703726

  1. Affine Vertex Operator Algebras and Modular Linear Differential Equations

    NASA Astrophysics Data System (ADS)

    Arike, Yusuke; Kaneko, Masanobu; Nagatomo, Kiyokazu; Sakai, Yuichi

    2016-05-01

    In this paper, we list all affine vertex operator algebras of positive integral levels whose dimensions of spaces of characters are at most 5 and show that a basis of the space of characters of each affine vertex operator algebra in the list gives a fundamental system of solutions of a modular linear differential equation. Further, we determine the dimensions of the spaces of characters of affine vertex operator algebras whose numbers of inequivalent simple modules are not exceeding 20.

  2. Novel thermo-responsive fucose binding ligands for glycoprotein purification by affinity precipitation.

    PubMed

    Arnold, Lindsay; Chen, Rachel

    2014-02-01

    Novel thermo-responsive affinity sugar binders were developed by fusing a bacterial fucose lectin with a thermo-responsive polypeptide. These designer affinity ligand fusions were produced using an Escherichia coli system capable of extracellular secretion of recombinant proteins and were isolated with a high recovery yield (95%) directly from growth medium by Inverse Temperature Cycling (ITC). With horse radish peroxidase (HRP) as a model protein, we demonstrate here that the designer thermo-responsive ligands are capable of interacting with glycans on a glycoprotein, a property that was used to develop a novel affinity precipitation method for glycoprotein purification. The method, requiring only simple process steps, affords full recovery of a target glycoprotein, and is effective at a target glycoprotein concentration as low as 1.4 pM in the presence of large amounts of contaminants. By developing other sugar binders in the similar fashion, the method should be highly useful for glycoprotein purification and detection. PMID:25271333

  3. Affinity Spaces, Apprenticeships, and Agency: Exploring Blogging Engagements in Family Spaces

    ERIC Educational Resources Information Center

    Lewis, Tisha Y.

    2014-01-01

    This empirical study examines the virtual exchanges between an African American mother and son's engagement in a blog. Using literacy as social practices as a framework, this study describes how blogging was developed and used to create affinity spaces, encouraged apprenticeship models, and shaped agentic roles within their family. Attention…

  4. High Affinity Iron Permease is Required for Virulence of Rhizopus oryzae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rhizopus oryzae is the most common cause of mucormycosis. Clinical and animal model data clearly demonstrate that the presence of elevated available serum iron predisposes the host to develop mucormycosis. The high affinity iron permease gene (rFTR1) is required for R. oryzae iron transport in iro...

  5. CALCULATION OF ELECTRON AFFINITIES OF POLYCYCLIC AROMATIC HYDROCARBONS AND SOVATION ENERGIES OF THEIR ANIONS

    EPA Science Inventory

    Electron affinities (EAs) and free energies for electron attachment have been calculated for 42 polynuclear aromatic hydrocarbons and related molecules by a variety of theoretical models, including Koopmans' theorem methods and the L1E method from differences in energy between th...

  6. Comparing binding site information to binding affinity reveals that Crp/DNA complexes have several distinct binding conformers

    PubMed Central

    Holmquist, Peter C.; Holmquist, Gerald P.; Summers, Michael L.

    2011-01-01

    We show that the cAMP receptor protein (Crp) binds to DNA as several different conformers. This situation has precluded discovering a high correlation between any sequence property and binding affinity for proteins that bend DNA. Experimentally quantified affinities of Synechocystis sp. PCC 6803 cAMP receptor protein (SyCrp1), the Escherichia coli Crp (EcCrp, also CAP) and DNA were analyzed to mathematically describe, and make human-readable, the relationship of DNA sequence and binding affinity in a given system. Here, sequence logos and weight matrices were built to model SyCrp1 binding sequences. Comparing the weight matrix model to binding affinity revealed several distinct binding conformations. These Crp/DNA conformations were asymmetrical (non-palindromic). PMID:21586590

  7. Antibody response and antibody affinity maturation in cats with experimental proliferative immune complex glomerulonephritis.

    PubMed

    Bishop, S A; Bailey, M; Lucke, V M; Stokes, C R

    1992-07-01

    An experimental model of proliferative glomerulonephritis (GN) in the cat, which closely resembles human proliferative forms of GN, has been used to study the role of antibody and antibody affinity in the development of immune complex-mediated renal disease. The serum IgG and IgM antibody response to antigen, average antibody affinity (avidity) and affinity heterogeneity of the IgG and IgM populations was assessed at varying times after commencement of chronic immunization with the antigen, human serum albumin (HSA), by enzyme immunoassay. Cats could be classified according to whether they were "low", "intermediate" or "high" IgG responders, by quantification of serum IgG values. Cats with the lowest serum IgG values failed to develop glomerulonephritis. However, there was no relationship between actual IgG values and the severity of the induced disease. In contrast to IgG, there was no division of cats into low or high IgM anti-HSA responders. Again, cats with the lowest IgM values failed to develop GN, but, more interestingly, a late, marked increase in serum IgM anti-HSA occurred only in cats that developed clinical signs of GN (anterior uveitis and nephrotic syndrome). Maturation of average, functional IgG affinity (avidity) for HSA following chronic immunization was clearly demonstrated for all cats. At the end of the experiment, all cats had IgG of high affinity for HSA and the average affinity heterogeneity of the IgG populations was less than in measurements taken earlier. Values of IgG affinity at the end of the experiment were very similar both in cats which developed GN and in those which remained clinically, biochemically and pathologically normal. In contrast to IgG antibody, some cats developed IgM of increased affinity, whilst others produced antibody of reduced affinity, following chronic immunization. There was no correlation between the development of disease and the production of either low or high affinity IgM antibody. Data indicated that an

  8. Different promoter affinities account for specificity in MYC-dependent gene regulation

    PubMed Central

    Lorenzin, Francesca; Benary, Uwe; Baluapuri, Apoorva; Walz, Susanne; Jung, Lisa Anna; von Eyss, Björn; Kisker, Caroline; Wolf, Jana; Eilers, Martin; Wolf, Elmar

    2016-01-01

    Enhanced expression of the MYC transcription factor is observed in the majority of tumors. Two seemingly conflicting models have been proposed for its function: one proposes that MYC enhances expression of all genes, while the other model suggests gene-specific regulation. Here, we have explored the hypothesis that specific gene expression profiles arise since promoters differ in affinity for MYC and high-affinity promoters are fully occupied by physiological levels of MYC. We determined cellular MYC levels and used RNA- and ChIP-sequencing to correlate promoter occupancy with gene expression at different concentrations of MYC. Mathematical modeling showed that binding affinities for interactions of MYC with DNA and with core promoter-bound factors, such as WDR5, are sufficient to explain promoter occupancies observed in vivo. Importantly, promoter affinity stratifies different biological processes that are regulated by MYC, explaining why tumor-specific MYC levels induce specific gene expression programs and alter defined biological properties of cells. DOI: http://dx.doi.org/10.7554/eLife.15161.001 PMID:27460974

  9. A Differential Dielectric Affinity Glucose Sensor

    PubMed Central

    Huang, Xian; Leduc, Charles; Ravussin, Yann; Li, Siqi; Davis, Erin; Song, Bing; Li, Dachao; Xu, Kexin; Accili, Domenico; Wang, Qian; Leibel, Rudolph; Lin, Qiao

    2013-01-01

    A continuous glucose monitor with a differential dielectric sensor implanted within the subcutaneous tissue that determines the glucose in the interstitial fluid is presented. The device, created using microelectromechanical systems (MEMS) technology, consists of sensing and reference modules that are identical in design and placed in close proximity. Each module contains a microchamber housing a pair of capacitive electrodes residing on the device substrate and embedded in a suspended, perforated polymer diaphragm. The microchambers, enclosed in semi-permeable membranes, are filled with either a polymer solution that has specific affinity to glucose or a glucose-insensitive reference solution. To accurately determine the glucose concentration, changes in the permittivity of the sensing and the reference solutions induced by changes in glucose concentration are measured differentially. In vitro characterization demonstrated the sensor capable of measuring glucose concentrations from 0 to 500 mg/dL with resolution and accuracy of ∼1.7 μg/dL and ∼1.74 mg/dL, respectively. In addition, device drift was reduced to 1.4% (uncontrolled environment) and 11% (5 °C of temperature variation) of that from non-differential measurements, indicating significant stability improvements. Preliminary animal testing demonstrated that the differential sensor accurately tracks glucose concentration in blood. This sensor can potentially be used clinically as a subcutaneously implanted continuous monitoring device in diabetic patients. PMID:24220675

  10. Affine gravity, Palatini formalism and charges

    NASA Astrophysics Data System (ADS)

    Katz, Joseph; Livshits, Gideon I.

    2011-12-01

    Affine gravity and the Palatini formalism contribute both to produce a simple and unique formula for calculating charges at spatial and null infinity for Lovelock type Lagrangians whose variational derivatives do not depend on second-order derivatives of the field components. The method is based on the covariant generalization due to Julia and Silva of the Regge-Teitelboim procedure that was used to define properly the mass in the classical formulation of Einstein's theory of gravity. Numerous applications reproduce standard results obtained by other secure but mostly specialized method like in ADM energy for asymptotically flat spacetimes and in Abbot and Deser for asymptotically de Sitter and anti-de Sitter spacetimes, both at spatial infinity. As a novel application we calculate the Bondi energy loss in five dimensional gravity, based on the asymptotic solution given by Tanabe et al. and obtain, as expected, the same result. We also give the for Einstein-Gauss-Bonnet gravity and find the superpotential for Lovelock theories of gravity when the number of dimensions tends to infinity with maximally symmetrical boundaries. The paper is written in standard component formalism.

  11. Divalent cation affinity sites in Paramecium aurelia.

    PubMed

    Fisher, G; Kaneshiro, E S; Peters, P D

    1976-05-01

    Sites with high calcium affinity in Paramecium aurelia were identified by high calcium (5 mM) fixation and electron microscope methods. Electron-opaque deposits were observed on the cytoplasmic side of surface membranes, particularly at the basal regions of cilia and trichocyst-pellicle fusion sites. Deposits were also observed on some smooth cytomembranes, within the axoneme of cilia, and on basal bodies. The divalent cations, Mg2+, Mn2+, Sr2+, Ni2+, Ba2+, and Zn2+, could be substituted for Ca2+ in the procedure. Deposits were larger with 5 mM Sr2+. Ba2+, and Mn2+ at ciliary transverse plates and the terminal plate of basal bodies. Microprobe analysis showed that Ca and C1 were concentrated within deposits. In some analyses, S and P were detected in deposits. Also, microprobe analysis of 5 mM Mn2+-fixed P. aurelia showed that those deposits were enriched in Mn and C1 and sometimes enriched in P. Deposits were seen only when the ciliates were actively swimming at the time of fixation. Locomotory mutants having defective membrane Ca-gating mechanisms and ciliates fixed while exhibiting ciliary reversal showed no obvious differences in deposition pattern and intensity. Possible correlations between electron-opaque deposits and the locations of intramembranous particles seen by freeze-fracture studied, as well as sites where fibrillar material associate with membranes are considered. The possibility that the action sites of calcium and other divalent cations were identified is discussed. PMID:1262398

  12. Multiplexed protein profiling by sequential affinity capture.

    PubMed

    Ayoglu, Burcu; Birgersson, Elin; Mezger, Anja; Nilsson, Mats; Uhlén, Mathias; Nilsson, Peter; Schwenk, Jochen M

    2016-04-01

    Antibody microarrays enable parallelized and miniaturized analysis of clinical samples, and have proven to provide novel insights for the analysis of different proteomes. However, there are concerns that the performance of such direct labeling and single antibody assays are prone to off-target binding due to the sample context. To improve selectivity and sensitivity while maintaining the possibility to conduct multiplexed protein profiling, we developed a multiplexed and semi-automated sequential capture assay. This novel bead-based procedure encompasses a first antigen capture, labeling of captured protein targets on magnetic particles, combinatorial target elution and a read-out by a secondary capture bead array. We demonstrate in a proof-of-concept setting that target detection via two sequential affinity interactions reduced off-target contribution, while lowered background and noise levels, improved correlation to clinical values compared to single binder assays. We also compared sensitivity levels with single binder and classical sandwich assays, explored the possibility for DNA-based signal amplification, and demonstrate the applicability of the dual capture bead-based antibody microarray for biomarker analysis. Hence, the described concept enhances the possibilities for antibody array assays to be utilized for protein profiling in body fluids and beyond. PMID:26935855

  13. Affinity of guanosine derivatives for polycytidylate revisited

    NASA Technical Reports Server (NTRS)

    Kanavarioti, A.; Hurley, T. B.; Baird, E. E.

    1995-01-01

    Evidence is presented for complexation of guanosine 5'-monophosphate 2-methylimidazolide (2-MeImpG) with polycytidylate (poly(C)) at pH 8.0 and 23 degrees C in the presence of 1.0 M NaCl2 and 0.2 M MgCl2 in water. The association of 2-MeImpG with poly(C) was investigated using UV-vis spectroscopy as well as by monitoring the kinetics of the nucleophilic substitution reaction of the imidazole moiety by amines. The results of both methods are consistent with moderately strong poly(C) 2-MeImpG complexation and the spectrophotometric measurements allowed the construction of a binding isotherm with a concentration of 2-MeImpG equal to 5.55 +/- 0.15 mM at half occupancy. UV spectroscopy was employed to establish the binding of other guanosine derivatives on poly(C). These derivatives are guanosine 5'-monophosphate (5'GMP), guanosine 5'-monophosphate imidazolide (ImpG), and guanosine 5'-monophosphate morpholidate (morpG). Within experimental error these guanosine derivatives exhibit the same affinity for poly(C) as 2-MeImpG.

  14. Banach frames in the affine synthesis problem

    NASA Astrophysics Data System (ADS)

    Terekhin, Pavel A.

    2009-10-01

    We consider the problem of representing functions f\\in L^p(\\mathbb R^d) by a series in elements of the affine system \\displaystyle \\psi_{j,k}(x)=\\lvert\\det a_j\\rvert^{1/2}\\psi(a_jx-bk), \\qquad j\\in\\mathbb N, \\quad k\\in\\mathbb Z^d. The corresponding representation theorems are established on the basis of the frame inequalities \\displaystyle A\\Vert g\\Vert _q\\le\\Vert\\{(g,\\psi_{j,k})\\}\\Vert _Y\\le B\\Vert g\\Vert _q for the Fourier coefficients \\displaystyle(g,\\psi_{j,k})=\\int_{\\mathbb R^d}g(x)\\psi_{j,k}(x)\\,dx of functions g\\in L^q(\\mathbb R^d), 1/p+1/q=1, where {\\Vert\\cdot\\Vert}_Y is the norm in some Banach space of number families \\{y_{j,k}\\} and 0 are constants. In particular, it is proved that if the integral of a function \\psi\\in L^1\\cap L^p(\\mathbb R^d), 1, is nonzero, so \\displaystyle\\int_{\\mathbb R^d}\\psi(x)\\,dx\

  15. FAST TRACK COMMUNICATION: Affine constellations without mutually unbiased counterparts

    NASA Astrophysics Data System (ADS)

    Weigert, Stefan; Durt, Thomas

    2010-10-01

    It has been conjectured that a complete set of mutually unbiased bases in a space of dimension d exists if and only if there is an affine plane of order d. We introduce affine constellations and compare their existence properties with those of mutually unbiased constellations. The observed discrepancies make a deeper relation between the two existence problems unlikely.

  16. The Study of Affinity-Seeking in an Organizational Setting.

    ERIC Educational Resources Information Center

    Flath, Dominic B.

    This study investigated the relationship between supervisors' use of Bell and Daly's affinity-seeking strategies and their impact on employee satisfaction. Results indicated that 16 of the 25 affinity-seeking strategies were positively correlated with a subordinate's perception of supervisor credibility. Results also indicated that a supervisor's…

  17. A minimax approach to spatial estimation using affinity matrices

    NASA Technical Reports Server (NTRS)

    Morris, C. N.

    1983-01-01

    Estimates made in the plane to improve on noisy unbiased estimates were combined. Only a small fraction of points in a giant grid were used to do this, those that are most like a given point. A component of this process defining an affinity matrix of values, indicating which points are relevant to others. Minimax rules are shown to be based on affinity matrices.

  18. Striving for Empathy: Affinities, Alliances and Peer Sexuality Educators

    ERIC Educational Resources Information Center

    Fields, Jessica; Copp, Martha

    2015-01-01

    Peer sexuality educators' accounts of their work reveal two approaches to empathy with their students: affinity and alliance. "Affinity-based empathy" rests on the idea that the more commonalities sexuality educators and students share (or perceive they share), the more they will be able to empathise with one another, while…

  19. Conformational kinetics reveals affinities of protein conformational states

    PubMed Central

    Daniels, Kyle G.; Suo, Yang; Oas, Terrence G.

    2015-01-01

    Most biological reactions rely on interplay between binding and changes in both macromolecular structure and dynamics. Practical understanding of this interplay requires detection of critical intermediates and determination of their binding and conformational characteristics. However, many of these species are only transiently present and they have often been overlooked in mechanistic studies of reactions that couple binding to conformational change. We monitored the kinetics of ligand-induced conformational changes in a small protein using six different ligands. We analyzed the kinetic data to simultaneously determine both binding affinities for the conformational states and the rate constants of conformational change. The approach we used is sufficiently robust to determine the affinities of three conformational states and detect even modest differences in the protein’s affinities for relatively similar ligands. Ligand binding favors higher-affinity conformational states by increasing forward conformational rate constants and/or decreasing reverse conformational rate constants. The amounts by which forward rate constants increase and reverse rate constants decrease are proportional to the ratio of affinities of the conformational states. We also show that both the affinity ratio and another parameter, which quantifies the changes in conformational rate constants upon ligand binding, are strong determinants of the mechanism (conformational selection and/or induced fit) of molecular recognition. Our results highlight the utility of analyzing the kinetics of conformational changes to determine affinities that cannot be determined from equilibrium experiments. Most importantly, they demonstrate an inextricable link between conformational dynamics and the binding affinities of conformational states. PMID:26162682

  20. Binding affinities of CRBPI and CRBPII for 9-cis-retinoids

    PubMed Central

    Kane, Maureen A.; Bright, Frank V.; Napoli, Joseph L.

    2014-01-01

    Background Cellular retinol binding-protein I (CRBPI) and cellular retinol binding-protein II (CRBPII) serve as intracellular retinoid chaperones that bind retinol and retinal with high affinity and facilitate substrate delivery to select enzymes that catalyze retinoic acid (RA) and retinyl ester biosynthesis. Recently, 9-cis-RA has been identified in vivo in the pancreas, where it contributes to regulating glucose-stimulated insulin secretion. In vitro, 9-cis-RA activates RXR (retinoid×receptors), which serve as therapeutic targets for treating cancer and metabolic diseases. Binding affinities and structure–function relationships have been well characterized for CRBPI and CRBPII with all-trans-retinoids, but not for 9-cis-retinoids. This study extended current knowledge by establishing binding affinities for CRBPI and CRBPII with 9-cis-retinoids. Methods We have determined apparent dissociation constants, Kd′, through monitoring binding of 9-cis-retinol, 9-cis-retinal, and 9-cis-RA with CRBPI and CRBPII by fluorescence spectroscopy, and analyzing the data with non-linear regression. We compared these data to the data we obtained for all-trans- and 13-cis-retinoids under identical conditions. Results CRBPI and CRBPII, respectively, bind 9-cis-retinol ( Kd′, 11 nM and 68 nM) and 9-cis-retinal ( Kd′, 8 nM and 5 nM) with high affinity. No significant 9-cis-RA binding was observed with CRBPI or CRBPII. Conclusions CRBPI and CRBPII bind 9-cis-retinol and 9-cis-retinal with high affinities, albeit with affinities somewhat lower than for all-trans-retinol and all-trans-retinal. General significance These data provide further insight into structure–binding relationships of cellular retinol binding-proteins and are consistent with a model of 9-cis-RA biosynthesis that involves chaperoned delivery of 9-cis-retinoids to enzymes that recognize retinoid binding-proteins. PMID:21382444

  1. Concurrent low- and high-affinity sulfate reduction kinetics in marine sediment

    NASA Astrophysics Data System (ADS)

    Harder Tarpgaard, Irene; Røy, Hans; Jørgensen, Bo Barker

    Bacterial sulfate reduction in marine sediments generally occurs in the presence of high millimolar concentrations of sulfate. Published data indicate that low sulfate concentrations may limit sulfate reduction rates below 0.2-2 mM. Yet, high sulfate reduction rates occur in the 1-100 μM range in freshwater sediments and at the sulfate-methane transition in marine sediments. Through a combination of 35S-tracer experiments, including initial velocity experiments and time course experiments, we searched for different sulfate affinities in the mixed community of sulfate reducers in a marine sediment. We supported the radiotracer experiments with a highly sensitive ion chromatographic technique for sulfate with a detection limit of 0.15 μM SO 42- in marine pore water. Our results showed that high and low affinities for sulfate co-occur and that the applied experimental approach may determine the observed apparent half saturation constant, Km. Our experimental and model data both show that sulfate reduction in the studied marine sediment could be explained by two dominating affinities for sulfate: a low affinity with a mean half saturation constant, Km, of 430 μM SO 42- and a high affinity with a mean Km of 2.6 μM SO 42-. The high-affinity sulfate reduction was thermodynamically un-constrained down to <1 μM SO 42-, both in our experiments and under in situ conditions. The reduction of radio-labeled sulfate was partly reversible due to concurrent re-oxidation of sulfide by Fe(III) and possibly due to a reversibility of the enzymatic pathway of sulfate reduction. A literature survey of apparent Km values for sediments and pure cultures is presented and discussed.

  2. Optimal T-cell receptor affinity for inducing autoimmunity.

    PubMed

    Koehli, Sabrina; Naeher, Dieter; Galati-Fournier, Virginie; Zehn, Dietmar; Palmer, Ed

    2014-12-01

    T-cell receptor affinity for self-antigen has an important role in establishing self-tolerance. Three transgenic mouse strains expressing antigens of variable affinity for the OVA transgenic-I T-cell receptor were generated to address how TCR affinity affects the efficiency of negative selection, the ability to prime an autoimmune response, and the elimination of the relevant target cell. Mice expressing antigens with an affinity just above the negative selection threshold exhibited the highest risk of developing experimental autoimmune diabetes. The data demonstrate that close to the affinity threshold for negative selection, sufficient numbers of self-reactive T cells escape deletion and create an increased risk for the development of autoimmunity. PMID:25411315

  3. Affinity Monolith-Integrated Microchips for Protein Purification and Concentration.

    PubMed

    Gao, Changlu; Sun, Xiuhua; Wang, Huaixin; Qiao, Wei; Hu, Bo

    2016-01-01

    Affinity chromatography is a valuable method to purify and concentrate minute amount of proteins. Monoliths with epoxy groups for affinity immobilization were prepared by direct in-situ photopolymerization of glycidyl methacrylate and ethylene glycol dimethacrylate in porogenic solvents consisting of 1-dodecanol and cyclohexanol. By integrating affinity monoliths onto a microfluidic system, targeted biomolecules can be captured and retained on affinity column, while other biomolecules having no specific interactions toward the immobilized ligands flow through the microchannel. Therefore, proteins which remain on the affinity column are purified and concentrated, and then eluted by appropriate solutions and finally, separated by microchip capillary electrophoresis. This integrated microfluidic device has been applied to the purification and separation of specific proteins (FITC-labeled human serum albumin and IgG) in a mixture. PMID:27473483

  4. Structural Change in the Dynein Stalk Region Associated with Two Different Affinities for the Microtubule.

    PubMed

    Nishikawa, Yosuke; Inatomi, Momoko; Iwasaki, Haruka; Kurisu, Genji

    2016-05-01

    Dynein is a large microtubule-based motor complex that requires tight coupling of intra-molecular ATP hydrolysis with the generation of mechanical force and track-binding activity. However, the microtubule-binding domain is structurally separated by about 15nm from the nucleotide-binding sites by a coiled-coil stalk. Thus, long-range two-way communication is necessary for coordination between the catalytic cycle of ATP hydrolysis and dynein's track-binding affinities. To investigate the structural changes that occur in the dynein stalk region to produce two different microtubule affinities, here we improve the resolution limit of the previously reported structure of the entire stalk region and we investigate structural changes in the dynein stalk and strut/buttress regions by comparing currently available X-ray structures. In the light of recent crystal structures, the basis of the transition from the low-affinity to the high-affinity coiled-coil registry is discussed. A concerted movement model previously reported by Carter and Vale is modified more specifically, and we proposed it as the open zipper model. PMID:26585405

  5. Chasing polys: Interdisciplinary affinity and its connection to physics identity

    NASA Astrophysics Data System (ADS)

    Scott, Tyler D.

    This research is based on two motivations that merge by means of the frameworks of interdisciplinary affinity and physics identity. First, a goal of education is to develop interdisciplinary abilities in students' thinking and work. But an often ignored factor is students interests and beliefs about being interdisciplinary. Thus, this work develops and uses a framework called interdisciplinary affinity. It encompasses students interests in making connections across disciplines and their beliefs about their abilities to make those connections. The second motivation of this research is to better understand how to engage more students with physics. Physics identity describes how a student sees themselves in relation to physics. By understanding how physics identity is developed, researchers and educators can identify factors that increase interest and engagement in physics classrooms. Therefore, physics identity was used in conjunction with interdisciplinary affinity. Using a mixed methods approach, this research used quantitative data to identify the relationships interdisciplinary affinity has with physics identity and the physics classroom. These connections were explored in more detail using a case study of three students in a high school physics class. Results showed significant and positive relationships between interdisciplinary affinity and physics identity, including the individual interest and recognition components of identity. It also identified characteristics of physics classrooms that had a significant, positive relationship with interdisciplinary affinity. The qualitative case study highlighted the importance of student interest to the relationship between interdisciplinary affinity and physics identity. It also identified interest and mastery orientation as key to understanding the link between interdisciplinary affinity and the physics classroom. These results are a positive sign that by understanding interdisciplinary affinity and physics identity

  6. Genetic affinities of central China populations.

    PubMed

    Zhou, H Y; Wang, H W; Tan, S N; Chen, Y; Wang, W L; Tao, H X; Yin, Z C; Zou, Y H; Ouyang, S M; Ni, B

    2014-01-01

    Hunan locates in the south-central part of China, to the south of the middle reaches of the Yangtze River and south of Lake Dongting. According to the historical records, the peopling of Hunan by modern human ancestors can ascend to 40 thousand years ago. Thus, to trace the ancient maternal components can offer further insight into the origin of south-central China. In this study, we investigated the mitochondrial DNA of 114 individuals from Hunan Province (including 34 Han, 40 Tujia and 40 Miao). Hypervariable regions I and II of the mtDNA control region were sequenced, and the relative diagnostic variations in coding region according to the updated worldwide phylogeny tree were selected and typed by restriction fragment length polymorphism analysis or direct sequencing. All individuals were classified into specific (sub)haplogroups. By comparison with the surrounding populations, southern China-prevalent haplogroups were detected with relative higher frequency in the Tujia and Miao ethnic populations, such as haplogroup B, with more than 20%, lacking in the Han population, which illustrated its southern origin characters. In addition, we also detected northern of East Asia prevalent haplogroups with a relative higher frequency in Tujia populations than in the Miao and Yao ethnic groups, implying a gene flow from Han populations. However, the language-clustering tendency was supported by our principal component analysis and further genetic estimation results. Han and ethnic groups in central China exhibited specific ancestors related to their closer language affinity, although there was extensively genetic admixture between Han and ethnic groups. PMID:24615027

  7. Selectively Promiscuous Opioid Ligands: Discovery of High Affinity/Low Efficacy Opioid Ligands with Substantial Nociceptin Opioid Peptide Receptor Affinity

    PubMed Central

    2015-01-01

    Emerging clinical and preclinical evidence suggests that a compound displaying high affinity for μ, κ, and δ opioid (MOP, KOP, and DOP) receptors and antagonist activity at each, coupled with moderate affinity and efficacy at nociceptin opioid peptide (NOP) receptors will have utility as a relapse prevention agent for multiple types of drug abuse. Members of the orvinol family of opioid ligands have the desired affinity profile but have typically displayed substantial efficacy at MOP and or KOP receptors. In this study it is shown that a phenyl ring analogue (1d) of buprenorphine displays the desired profile in vitro with high, nonselective affinity for the MOP, KOP, and DOP receptors coupled with moderate affinity for NOP receptors. In vivo, 1d lacked any opioid agonist activity and was an antagonist of both the MOP receptor agonist morphine and the KOP receptor agonist ethylketocyclazocine, confirming the desired opioid receptor profile in vivo. PMID:24761755

  8. Nano-Bonding of Silicon Oxides-based surfaces at Low Temperature: Bonding Interphase Modeling via Molecular Dynamics and Characterization of Bonding Surfaces Topography, Hydro-affinity and Free Energy

    NASA Astrophysics Data System (ADS)

    Whaley, Shawn D.

    In this work, a new method, "Nanobonding(TM)" [1,2] is conceived and researched to bond Si-based surfaces, via nucleation and growth of a 2 D silicon oxide SiOxHx interphase connecting the surfaces at the nanoscale across macroscopic domains. Nanobonding cross-bridges two smooth surfaces put into mechanical contact in an O2/H 2O mixed ambient below T ≤200 °C via arrays of SiOxH x molecules connecting into a continuous macroscopic bonding interphase. Nano-scale surface planarization via wet chemical processing and new spin technology are compared via Tapping Mode Atomic Force Microscopy (TMAFM) , before and after nano-bonding. Nanobonding uses precursor phases, 2D nano-films of beta-cristobalite (beta-c) SiO2, nucleated on Si(100) via the Herbots-Atluri (H-A) method [1]. beta-c SiO2 on Si(100) is ordered and flat with atomic terraces over 20 nm wide, well above 2 nm found in native oxides. When contacted with SiO2 this ultra-smooth nanophase can nucleate and grow domains with cross-bridging molecular strands of hydroxylated SiOx, instead of point contacts. The high density of molecular bonds across extended terraces forms a strong bond between Si-based substrates, nano- bonding [2] the Si and silica. A new model of beta-cristobalite SiO2 with its <110> axis aligned along Si[100] direction is simulated via ab-initio methods in a nano-bonded stack with beta-c SiO2 in contact with amorphous SiO2 (a-SiO2), modelling cross-bridging molecular bonds between beta-c SiO2 on Si(100) and a-SiO2 as during nanobonding. Computed total energies are compared with those found for Si(100) and a-SiO2 and show that the presence of two lattice cells of !-c SiO2 on Si(100) and a-SiO2 lowers energy when compared to Si(100)/ a-SiO 2 Shadow cone calculations on three models of beta-c SiO2 on Si(100) are compared with Ion Beam Analysis of H-A processed Si(100). Total surface energy measurements via 3 liquid contact angle analysis of Si(100) after H-A method processing are also compared

  9. Changes in Binding of [(123)I]CLINDE, a High-Affinity Translocator Protein 18 kDa (TSPO) Selective Radioligand in a Rat Model of Traumatic Brain Injury.

    PubMed

    Donat, Cornelius K; Gaber, Khaled; Meixensberger, Jürgen; Brust, Peter; Pinborg, Lars H; Hansen, Henrik H; Mikkelsen, Jens D

    2016-06-01

    After traumatic brain injury (TBI), secondary injuries develop, including neuroinflammatory processes that contribute to long-lasting impairments. These secondary injuries represent potential targets for treatment and diagnostics. The translocator protein 18 kDa (TSPO) is expressed in activated microglia cells and upregulated in response to brain injury and therefore a potential biomarker of the neuroinflammatory processes. Second-generation radioligands of TSPO, such as [(123)I]CLINDE, have a higher signal-to-noise ratio as the prototype ligand PK11195. [(123)I]CLINDE has been employed in human studies using single-photon emission computed tomography to image the neuroinflammatory response after stroke. In this study, we used the same tracer in a rat model of TBI to determine changes in TSPO expression. Adult Sprague-Dawley rats were subjected to moderate controlled cortical impact injury and sacrificed at 6, 24, 72 h and 28 days post surgery. TSPO expression was assessed in brain sections employing [(123)I]CLINDE in vitro autoradiography. From 24 h to 28 days post surgery, injured animals exhibited a marked and time-dependent increase in [(123)I]CLINDE binding in the ipsilateral motor, somatosensory and parietal cortex, as well as in the hippocampus and thalamus. Interestingly, binding was also significantly elevated in the contralateral M1 motor cortex following TBI. Craniotomy without TBI caused a less marked increase in [(123)I]CLINDE binding, restricted to the ipsilateral hemisphere. Radioligand binding was consistent with an increase in TSPO mRNA expression and CD11b immunoreactivity at the contusion site. This study demonstrates the applicability of [(123)I]CLINDE for detailed regional and quantitative assessment of glial activity in experimental models of TBI. PMID:26969181

  10. Tetrahydroprotoberberine alkaloids with dopamine and σ receptor affinity.

    PubMed

    Gadhiya, Satishkumar; Madapa, Sudharshan; Kurtzman, Thomas; Alberts, Ian L; Ramsey, Steven; Pillarsetty, Nagavara-Kishore; Kalidindi, Teja; Harding, Wayne W

    2016-05-01

    Two series of analogues of the tetrahydroprotoberberine (THPB) alkaloid (±)-stepholidine that (a) contain various alkoxy substituents at the C10 position and, (b) were de-rigidified with respect to (±)-stepholidine, were synthesized and evaluated for affinity at dopamine and σ receptors in order to evaluate effects on D3 and σ2 receptor affinity and selectivity. Small n-alkoxy groups are best tolerated by D3 and σ2 receptors. Among all compounds tested, C10 methoxy and ethoxy analogues (10 and 11 respectively) displayed the highest affinity for σ2 receptors as well as σ2 versus σ1 selectivity and also showed the highest D3 receptor affinity. De-rigidification of stepholidine resulted in decreased affinity at all receptors evaluated; thus the tetracyclic THPB framework is advantageous for affinity at dopamine and σ receptors. Docking of the C10 analogues at the D3 receptor, suggest that an ionic interaction between the protonated nitrogen atom and Asp110, a H-bond interaction between the C2 phenol and Ser192, a H-bond interaction between the C10 phenol and Cys181 as well as hydrophobic interactions of the aryl rings to Phe106 and Phe345, are critical for high affinity of the compounds. PMID:27032890

  11. Analysis of biomolecular interactions using affinity microcolumns: A review

    PubMed Central

    Zheng, Xiwei; Li, Zhao; Beeram, Sandya; Podariu, Maria; Matsuda, Ryan; Pfaunmiller, Erika L.; White, Christopher J.; Carter, NaTasha; Hage, David S.

    2014-01-01

    Affinity chromatography has become an important tool for characterizing biomolecular interactions. The use of affinity microcolumns, which contain immobilized binding agents and have volumes in the mid-to-low microliter range, has received particular attention in recent years. Potential advantages of affinity microcolumns include the many analysis and detection formats that can be used with these columns, as well as the need for only small amounts of supports and immobilized binding agents. This review examines how affinity microcolumns have been used to examine biomolecular interactions. Both capillary-based microcolumns and short microcolumns are considered. The use of affinity microcolumns with zonal elution and frontal analysis methods are discussed. The techniques of peak decay analysis, ultrafast affinity extraction, split-peak analysis, and band-broadening studies are also explored. The principles of these methods are examined and various applications are provided to illustrate the use of these methods with affinity microcolumns. It is shown how these techniques can be utilized to provide information on the binding strength and kinetics of an interaction, as well as on the number and types of binding sites. It is further demonstrated how information on competition or displacement effects can be obtained by these methods. PMID:24572459

  12. Analysis of biomolecular interactions using affinity microcolumns: a review.

    PubMed

    Zheng, Xiwei; Li, Zhao; Beeram, Sandya; Podariu, Maria; Matsuda, Ryan; Pfaunmiller, Erika L; White, Christopher J; Carter, NaTasha; Hage, David S

    2014-10-01

    Affinity chromatography has become an important tool for characterizing biomolecular interactions. The use of affinity microcolumns, which contain immobilized binding agents and have volumes in the mid-to-low microliter range, has received particular attention in recent years. Potential advantages of affinity microcolumns include the many analysis and detection formats that can be used with these columns, as well as the need for only small amounts of supports and immobilized binding agents. This review examines how affinity microcolumns have been used to examine biomolecular interactions. Both capillary-based microcolumns and short microcolumns are considered. The use of affinity microcolumns with zonal elution and frontal analysis methods are discussed. The techniques of peak decay analysis, ultrafast affinity extraction, split-peak analysis, and band-broadening studies are also explored. The principles of these methods are examined and various applications are provided to illustrate the use of these methods with affinity microcolumns. It is shown how these techniques can be utilized to provide information on the binding strength and kinetics of an interaction, as well as on the number and types of binding sites. It is further demonstrated how information on competition or displacement effects can be obtained by these methods. PMID:24572459

  13. Methodological study of affine transformations of gene expression data with proposed robust non-parametric multi-dimensional normalization method

    PubMed Central

    Bengtsson, Henrik; Hössjer, Ola

    2006-01-01

    Background Low-level processing and normalization of microarray data are most important steps in microarray analysis, which have profound impact on downstream analysis. Multiple methods have been suggested to date, but it is not clear which is the best. It is therefore important to further study the different normalization methods in detail and the nature of microarray data in general. Results A methodological study of affine models for gene expression data is carried out. Focus is on two-channel comparative studies, but the findings generalize also to single- and multi-channel data. The discussion applies to spotted as well as in-situ synthesized microarray data. Existing normalization methods such as curve-fit ("lowess") normalization, parallel and perpendicular translation normalization, and quantile normalization, but also dye-swap normalization are revisited in the light of the affine model and their strengths and weaknesses are investigated in this context. As a direct result from this study, we propose a robust non-parametric multi-dimensional affine normalization method, which can be applied to any number of microarrays with any number of channels either individually or all at once. A high-quality cDNA microarray data set with spike-in controls is used to demonstrate the power of the affine model and the proposed normalization method. Conclusion We find that an affine model can explain non-linear intensity-dependent systematic effects in observed log-ratios. Affine normalization removes such artifacts for non-differentially expressed genes and assures that symmetry between negative and positive log-ratios is obtained, which is fundamental when identifying differentially expressed genes. In addition, affine normalization makes the empirical distributions in different channels more equal, which is the purpose of quantile normalization, and may also explain why dye-swap normalization works or fails. All methods are made available in the aroma package, which is

  14. Point mutation increases a form of the NK1 receptor with high affinity for neurokinin A and B and septide

    PubMed Central

    Ciucci, Alessandra; Palma, Carla; Manzini, Stefano; Werge, Thomas M

    1998-01-01

    The binding modalities of substance P and neurokinin A on the wild type and Gly166 to-Cys mutant NK1 receptors expressed on CHO cells were investigated in homologous and heterologous binding experiments using both radiolabelled substance P and neurokinin A.On the wild type NK1 receptor NKA displaces radiolabelled substance P with very low apparent affinity, despite its high-affinity binding constant (determined in homologous binding experiments). The Gly166 to-Cys substitution in the NK1 tachykinin receptor greatly enhances the apparent affinity of neurokinin A in competition for radiolabelled substance P, but it does not change the binding constant of neurokinin A. The mutation, thereby, eliminates the discrepancy between the low apparent affinity and the high binding constant of neurokinin A.On the wild type receptor the binding capacity of neurokinin A is significantly smaller than that of substance P. In contrast, the two tachykinins bind to approximately the same number of sites on the mutant receptor.Simultaneous mass action law analysis of binding data in which multiple radioligands were employed in parallel demonstrated that a one-site model was unable to accommodate all the experimental data, whereas a two-site model provided a dramatically better description.These two receptor-sites display equally high affinity for substance P, while neurokinin A strongly discriminates between a high and a low affinity component. The binding affinities of neurokinin A are not affected by the mutation, which instead specifically alters the distribution between receptor sites in favour of a high affinity neurokinin A binding form.The low apparent affinity and binding capacity of neurokinin A on the wild type receptor results from neurokinin A binding with high affinity only to a fraction of the sites labelled by substance P. The mutation increases the proportion of this site, and consequently enhances the apparent affinity and binding capacity of neurokinin A.The binding

  15. Predicting direct protein interactions from affinity purification mass spectrometry data

    PubMed Central

    2010-01-01

    Background Affinity purification followed by mass spectrometry identification (AP-MS) is an increasingly popular approach to observe protein-protein interactions (PPI) in vivo. One drawback of AP-MS, however, is that it is prone to detecting indirect interactions mixed with direct physical interactions. Therefore, the ability to distinguish direct interactions from indirect ones is of much interest. Results We first propose a simple probabilistic model for the interactions captured by AP-MS experiments, under which the problem of separating direct interactions from indirect ones is formulated. Then, given idealized quantitative AP-MS data, we study the problem of identifying the most likely set of direct interactions that produced the observed data. We address this challenging graph theoretical problem by first characterizing signatures that can identify weakly connected nodes as well as dense regions of the network. The rest of the direct PPI network is then inferred using a genetic algorithm. Our algorithm shows good performance on both simulated and biological networks with very high sensitivity and specificity. Then the algorithm is used to predict direct interactions from a set of AP-MS PPI data from yeast, and its performance is measured against a high-quality interaction dataset. Conclusions As the sensitivity of AP-MS pipeline improves, the fraction of indirect interactions detected will also increase, thereby making the ability to distinguish them even more desirable. Despite the simplicity of our model for indirect interactions, our method provides a good performance on the test networks. PMID:21034440

  16. Predicting the relative binding affinity of mineralocorticoid receptor antagonists by density functional methods.

    PubMed

    Roos, Katarina; Hogner, Anders; Ogg, Derek; Packer, Martin J; Hansson, Eva; Granberg, Kenneth L; Evertsson, Emma; Nordqvist, Anneli

    2015-12-01

    In drug discovery, prediction of binding affinity ahead of synthesis to aid compound prioritization is still hampered by the low throughput of the more accurate methods and the lack of general pertinence of one method that fits all systems. Here we show the applicability of a method based on density functional theory using core fragments and a protein model with only the first shell residues surrounding the core, to predict relative binding affinity of a matched series of mineralocorticoid receptor (MR) antagonists. Antagonists of MR are used for treatment of chronic heart failure and hypertension. Marketed MR antagonists, spironolactone and eplerenone, are also believed to be highly efficacious in treatment of chronic kidney disease in diabetes patients, but is contra-indicated due to the increased risk for hyperkalemia. These findings and a significant unmet medical need among patients with chronic kidney disease continues to stimulate efforts in the discovery of new MR antagonist with maintained efficacy but low or no risk for hyperkalemia. Applied on a matched series of MR antagonists the quantum mechanical based method gave an R(2) = 0.76 for the experimental lipophilic ligand efficiency versus relative predicted binding affinity calculated with the M06-2X functional in gas phase and an R(2) = 0.64 for experimental binding affinity versus relative predicted binding affinity calculated with the M06-2X functional including an implicit solvation model. The quantum mechanical approach using core fragments was compared to free energy perturbation calculations using the full sized compound structures. PMID:26572910

  17. General Anesthetics Predicted to Block the GLIC Pore with Micromolar Affinity

    PubMed Central

    Eckenhoff, Roderic G.; Klein, Michael L.; Brannigan, Grace

    2012-01-01

    Although general anesthetics are known to modulate the activity of ligand-gated ion channels in the Cys-loop superfamily, there is at present neither consensus on the underlying mechanisms, nor predictive models of this modulation. Viable models need to offer quantitative assessment of the relative importance of several identified anesthetic binding sites. However, to date, precise affinity data for individual sites has been challenging to obtain by biophysical means. Here, the likely role of pore block inhibition by the general anesthetics isoflurane and propofol of the prokaryotic pentameric channel GLIC is investigated by molecular simulations. Microscopic affinities are calculated for both single and double occupancy binding of isoflurane and propofol to the GLIC pore. Computations are carried out for an open-pore conformation in which the pore is restrained to crystallographic radius, and a closed-pore conformation that results from unrestrained molecular dynamics equilibration of the structure. The GLIC pore is predicted to be blocked at the micromolar concentrations for which inhibition by isofluorane and propofol is observed experimentally. Calculated affinities suggest that pore block by propofol occurs at signifcantly lower concentrations than those for which inhibition is observed: we argue that this discrepancy may result from binding of propofol to an allosteric site recently identified by X-ray crystallography, which may cause a competing gain-of-function effect. Affinities of isoflurane and propofol to the allosteric site are also calculated, and shown to be 3 mM for isoflurane and for propofol; both anesthetics have a lower affinity for the allosteric site than for the unoccupied pore. PMID:22693438

  18. Structure-based affinity maturation of a chimeric anti-ricin antibody C4C13.

    PubMed

    Luo, Longlong; Luo, Qun; Guo, Leiming; Lv, Ming; Lin, Zhou; Geng, Jing; Li, Xinying; Li, Yan; Shen, Beifen; Qiao, Chunxia; Feng, Jiannan

    2014-01-01

    Ricin is a highly lethal toxin. Anti-ricin chimeric monoclonal antibody (mAb) C4C13 was prepared in our lab; however, its binding affinity was much weaker than that of the parent antibody 4C13. In this study, based on the computer-guided homology modeling and conformational optimization methods, the 3-D structure of C4C13 variable regions Fv was constructed and optimized. Using molecular docking and dynamics simulation methods, the 3-D complex structure of ricin and C4C13 Fv was obtained. Considering the orientation property, surface electrostatic distribution, residues chemical and physical character and intermolecular hydrogen bond, the binding mode and key residues were predicted. According to C4C13 Fv fragment and ricin complementary binding surface, electrostatic attraction periphery and van der Waals interaction interface, three mutants (i.e., M1 (N(H102)F, W(H103)Y); M2 (W(H103)Y) and M3 (R(L90)G)) were designed, in which M1 and M2 were predicted to possess higher antigen-binding activity than C4C13, while M3 was weaker. The relative affinity assays by ELISA showed that M1 and M2 mutations had higher affinity (9.6 and 18.3 nmol/L) than C4C13 (130 nmol/L) and M3 had weaker affinity (234.5 nmol/L) than C4C13. The results showed that the modeling complex structure of the antigen (ricin) and antibody (C4C13) is reasonable. Our work offered affinity maturated antibodies by site mutations, which were beneficial for valuable anti-ricin antibody design and preparation in future. PMID:23527922

  19. Strategies to guide the antibody affinity maturation process.

    PubMed

    Doria-Rose, Nicole A; Joyce, M Gordon

    2015-04-01

    Antibodies with protective activity are critical for vaccine efficacy. Affinity maturation increases antibody activity through multiple rounds of somatic hypermutation and selection in the germinal center. Identification of HIV-1 specific and influenza-specific antibody developmental pathways, as well as characterization of B cell and virus co-evolution in patients, has informed our understanding of antibody development. In order to counteract HIV-1 and influenza viral diversity, broadly neutralizing antibodies precisely target specific sites of vulnerability and require high levels of affinity maturation. We present immunization strategies that attempt to recapitulate these natural processes and guide the affinity maturation process. PMID:25913818

  20. Strategies to guide the antibody affinity maturation process

    PubMed Central

    Doria-Rose, Nicole A.; Joyce, M. Gordon

    2015-01-01

    Antibodies with protective activity are critical for vaccine efficacy. Affinity maturation increases antibody activity through multiple rounds of somatic hypermutation and selection in the germinal center. Identification of HIV-1 specific and influenza-specific antibody developmental pathways, as well as characterization of B cell and virus co-evolution in patients, has informed our understanding of antibody development. In order to counteract HIV-1 and Influenza viral diversity, broadly neutralizing antibodies precisely target specific sites of vulnerability and require high levels of affinity maturation. We present immunization strategies that attempt to recapitulate these natural processes and guide the affinity maturation process. PMID:25913818

  1. Affinity- and topology-dependent bound on current fluctuations

    NASA Astrophysics Data System (ADS)

    Pietzonka, Patrick; Barato, Andre C.; Seifert, Udo

    2016-08-01

    We provide a proof of a recently conjectured universal bound on current fluctuations in Markovian processes. This bound establishes a link between the fluctuations of an individual observable current, the cycle affinities driving the system into a non-equilibrium steady state, and the topology of the network. The proof is based on a decomposition of the network into independent cycles with both positive affinity and positive stationary cycle current. This formalism allows for a refinement of the bound for systems in equilibrium or with locally vanishing affinities.

  2. Affinity+: Semi-Structured Brainstorming on Large Displays

    SciTech Connect

    Burtner, Edwin R.; May, Richard A.; Scarberry, Randall E.; LaMothe, Ryan R.; Endert, Alexander

    2013-04-27

    Affinity diagraming is a powerful method for encouraging and capturing lateral thinking in a group environment. The Affinity+ Concept was designed to improve the collaborative brainstorm process through the use of large display surfaces in conjunction with mobile devices like smart phones and tablets. The system works by capturing the ideas digitally and allowing users to sort and group them on a large touch screen manually. Additionally, Affinity+ incorporates theme detection, topic clustering, and other processing algorithms that help bring structured analytic techniques to the process without requiring explicit leadership roles and other overhead typically involved in these activities.

  3. Multi-Graph Matching via Affinity Optimization with Graduated Consistency Regularization.

    PubMed

    Yan, Junchi; Cho, Minsu; Zha, Hongyuan; Yang, Xiaokang; Chu, Stephen M

    2016-06-01

    This paper addresses the problem of matching common node correspondences among multiple graphs referring to an identical or related structure. This multi-graph matching problem involves two correlated components: i) the local pairwise matching affinity across pairs of graphs; ii) the global matching consistency that measures the uniqueness of the pairwise matchings by different composition orders. Previous studies typically either enforce the matching consistency constraints in the beginning of an iterative optimization, which may propagate matching error both over iterations and across graph pairs; or separate affinity optimization and consistency enforcement into two steps. This paper is motivated by the observation that matching consistency can serve as a regularizer in the affinity objective function especially when the function is biased due to noises or inappropriate modeling. We propose composition-based multi-graph matching methods to incorporate the two aspects by optimizing the affinity score, meanwhile gradually infusing the consistency. We also propose two mechanisms to elicit the common inliers against outliers. Compelling results on synthetic and real images show the competency of our algorithms. PMID:26372208

  4. Manipulating the selection forces during affinity maturation to generate cross-reactive HIV antibodies

    PubMed Central

    Wang, Shenshen; Mata-Fink, Jordi; Kriegsman, Barry; Hanson, Melissa; Irvine, Darrell J.; Eisen, Herman N.; Burton, Dennis R.; Wittrup, K. Dane; Kardar, Mehran; Chakraborty, Arup K.

    2015-01-01

    Summary Generation of potent antibodies by a mutation-selection process called affinity maturation is a key component of effective immune responses. Antibodies that protect against highly mutable pathogens must neutralize diverse strains. Developing effective immunization strategies to drive their evolution requires understanding how affinity maturation happens in an enviroment where variants of the same antigen are present. We present an in silico model of affinity maturation driven by antigen variants which reveals that induction of cross-reactive antibodies often occurs with low probability because conflicting selection forces, imposed by different antigen variants, can frustrate affinity maturation. We describe how variables such as temporal pattern of antigen administration influence the outcome of this frustrated evolutionary process. Our calculations predict, and experiments in mice with variant gp120 constructs of the HIV envelope protein confirm, that sequential immunization with antigen variants is preferred over a cocktail for induction of cross-reactive antibodies focused on the shared CD4 binding site epitope. PMID:25662010

  5. Structural correlates of affinity in fetal versus adult endplate nicotinic receptors

    NASA Astrophysics Data System (ADS)

    Nayak, Tapan Kumar; Chakraborty, Srirupa; Zheng, Wenjun; Auerbach, Anthony

    2016-04-01

    Adult-type nicotinic acetylcholine receptors (AChRs) mediate signalling at mature neuromuscular junctions and fetal-type AChRs are necessary for proper synapse development. Each AChR has two neurotransmitter binding sites located at the interface of a principal and a complementary subunit. Although all agonist binding sites have the same core of five aromatic amino acids, the fetal site has ~30-fold higher affinity for the neurotransmitter ACh. Here we use molecular dynamics simulations of adult versus fetal homology models to identify complementary-subunit residues near the core that influence affinity, and use single-channel electrophysiology to corroborate the results. Four residues in combination determine adult versus fetal affinity. Simulations suggest that at lower-affinity sites, one of these unsettles the core directly and the others (in loop E) increase backbone flexibility to unlock a key, complementary tryptophan from the core. Swapping only four amino acids is necessary and sufficient to exchange function between adult and fetal AChRs.

  6. Structural correlates of affinity in fetal versus adult endplate nicotinic receptors

    PubMed Central

    Nayak, Tapan Kumar; Chakraborty, Srirupa; Zheng, Wenjun; Auerbach, Anthony

    2016-01-01

    Adult-type nicotinic acetylcholine receptors (AChRs) mediate signalling at mature neuromuscular junctions and fetal-type AChRs are necessary for proper synapse development. Each AChR has two neurotransmitter binding sites located at the interface of a principal and a complementary subunit. Although all agonist binding sites have the same core of five aromatic amino acids, the fetal site has ∼30-fold higher affinity for the neurotransmitter ACh. Here we use molecular dynamics simulations of adult versus fetal homology models to identify complementary-subunit residues near the core that influence affinity, and use single-channel electrophysiology to corroborate the results. Four residues in combination determine adult versus fetal affinity. Simulations suggest that at lower-affinity sites, one of these unsettles the core directly and the others (in loop E) increase backbone flexibility to unlock a key, complementary tryptophan from the core. Swapping only four amino acids is necessary and sufficient to exchange function between adult and fetal AChRs. PMID:27101778

  7. Structural correlates of affinity in fetal versus adult endplate nicotinic receptors.

    PubMed

    Nayak, Tapan Kumar; Chakraborty, Srirupa; Zheng, Wenjun; Auerbach, Anthony

    2016-01-01

    Adult-type nicotinic acetylcholine receptors (AChRs) mediate signalling at mature neuromuscular junctions and fetal-type AChRs are necessary for proper synapse development. Each AChR has two neurotransmitter binding sites located at the interface of a principal and a complementary subunit. Although all agonist binding sites have the same core of five aromatic amino acids, the fetal site has ∼30-fold higher affinity for the neurotransmitter ACh. Here we use molecular dynamics simulations of adult versus fetal homology models to identify complementary-subunit residues near the core that influence affinity, and use single-channel electrophysiology to corroborate the results. Four residues in combination determine adult versus fetal affinity. Simulations suggest that at lower-affinity sites, one of these unsettles the core directly and the others (in loop E) increase backbone flexibility to unlock a key, complementary tryptophan from the core. Swapping only four amino acids is necessary and sufficient to exchange function between adult and fetal AChRs. PMID:27101778

  8. Tension-compression asymmetry in the binding affinity of membrane-anchored receptors and ligands

    NASA Astrophysics Data System (ADS)

    Xu, Guang-Kui; Liu, Zishun; Feng, Xi-Qiao; Gao, Huajian

    2016-03-01

    Cell adhesion plays a crucial role in many biological processes of cells, e.g., immune responses, tissue morphogenesis, and stem cell differentiation. An essential problem in the molecular mechanism of cell adhesion is to characterize the binding affinity of membrane-anchored receptors and ligands under different physiological conditions. In this paper, a theoretical model is presented to study the binding affinity between a large number of anchored receptors and ligands under both tensile and compressive stresses, and corroborated by demonstrating excellent agreement with Monte Carlo simulations. It is shown that the binding affinity becomes lower as the magnitude of the applied stress increases, and drops to zero at a critical tensile or compressive stress. Interestingly, the critical compressive stress is found to be substantially smaller than the critical tensile stress for relatively long and flexible receptor-ligand complexes. This counterintuitive finding is explained by using the Euler instability theory of slender columns under compression. The tension-compression asymmetry in the binding affinity of anchored receptors and ligands depends subtly on the competition between the breaking and instability of their complexes. This study helps in understanding the role of mechanical forces in cell adhesion mediated by specific binding molecules.

  9. The low-affinity complex of cytochrome c and its peroxidase

    PubMed Central

    Van de Water, Karen; Sterckx, Yann G. J.; Volkov, Alexander N.

    2015-01-01

    The complex of yeast cytochrome c peroxidase and cytochrome c is a paradigm of the biological electron transfer (ET). Building on seven decades of research, two different models have been proposed to explain its functional redox activity. One postulates that the intermolecular ET occurs only in the dominant, high-affinity protein–protein orientation, while the other posits formation of an additional, low-affinity complex, which is much more active than the dominant one. Unlike the high-affinity interaction—extensively studied by X-ray crystallography and NMR spectroscopy—until now the binding of cytochrome c to the low-affinity site has not been observed directly, but inferred mainly from kinetics experiments. Here we report the structure of this elusive, weak protein complex and show that it consists of a dominant, inactive bound species and an ensemble of minor, ET-competent protein–protein orientations, which summarily account for the experimentally determined value of the ET rate constant. PMID:25944250

  10. Network-of-queues approach to B-cell-receptor affinity discrimination

    NASA Astrophysics Data System (ADS)

    Felizzi, Federico; Comoglio, Federico

    2012-06-01

    The immune system is one of the most complex signal processing machineries in biology. The adaptive immune system, consisting of B and T lymphocytes, is activated in response to a large spectrum of pathogen antigens. B cells recognize and bind the antigen through B-cell receptors (BCRs) and this is fundamental for B-cell activation. However, the system response is dependent on BCR-antigen affinity values that span several orders of magnitude. Moreover, the ability of the BCR to discriminate between affinities at the high end (e.g., 109M-1-1010M-1) challenges the formulation of a mathematical model able to robustly separate these affinity-dependent responses. Queuing theory enables the analysis of many related processes, such as those resulting from the stochasticity of protein binding and unbinding events. Here we define a network of queues, consisting of BCR early signaling states and transition rates related to the propensity of molecular aggregates to form or disassemble. By considering the family of marginal distributions of BCRs in a given signaling state, we report a significant separation (measured as Jensen-Shannon divergence) that arises from a broad spectrum of antigen affinities.

  11. Network-of-queues approach to B-cell-receptor affinity discrimination.

    PubMed

    Felizzi, Federico; Comoglio, Federico

    2012-06-01

    The immune system is one of the most complex signal processing machineries in biology. The adaptive immune system, consisting of B and T lymphocytes, is activated in response to a large spectrum of pathogen antigens. B cells recognize and bind the antigen through B-cell receptors (BCRs) and this is fundamental for B-cell activation. However, the system response is dependent on BCR-antigen affinity values that span several orders of magnitude. Moreover, the ability of the BCR to discriminate between affinities at the high end (e.g., 10^{9}M^{-1}-10^{10}M^{-1}) challenges the formulation of a mathematical model able to robustly separate these affinity-dependent responses. Queuing theory enables the analysis of many related processes, such as those resulting from the stochasticity of protein binding and unbinding events. Here we define a network of queues, consisting of BCR early signaling states and transition rates related to the propensity of molecular aggregates to form or disassemble. By considering the family of marginal distributions of BCRs in a given signaling state, we report a significant separation (measured as Jensen-Shannon divergence) that arises from a broad spectrum of antigen affinities. PMID:23005146

  12. Radioligand binding assays for high affinity binders in the presence of endogenous ligands

    SciTech Connect

    White, H.B. III; McGahan, T.

    1986-05-01

    Endogenous ligands complicate radioligand-binding assays of high-affinity binding proteins by obscuring binding sites or by diluting the labeled ligand. They have developed a mathematical model for such systems where structurally identical radioligand and endogenous ligand can be equilibrated on the binding site and bound radioligand measured. A double-reciprocal plot of bound radioligand, *L/sub B/, versus sample volume, V, yields a straight line. Introduction of scaling factors for sample dilution, F, and total radioligand available, *L/sub T/, produces a plot in which the x-intercept yields the endogenous ligand concentration, (L/sub T/); the slope is the reciprocal of the binding protein concentration, (P/sub T/)/sup -1/; and the y-intercept is the fractional saturation of the high-affinity binder, L/sub T//P/sub T/. This type of analysis has been applied to the assay of high-affinity biotin-binding proteins in egg yolk. Its use led to the detection of a second biotin-binding protein which is heat labile. The conceptual approach can be applied to the assay of other high-affinity binders.

  13. Blind prediction of SAMPL4 cucurbit[7]uril binding affinities with the mining minima method

    PubMed Central

    Muddana, Hari S.; Yin, Jian; Sapra, Neil V.; Fenley, Andrew T.; Gilson, Michael K.

    2014-01-01

    Accurate methods for predicting protein-ligand binding affinities are of central interest to computer-aided drug design for hit identification and lead optimization. Here, we used the mining minima (M2) method to predict cucurbit[7]uril binding affinities from the SAMPL4 blind prediction challenge. We tested two different energy models, an empirical classical force field, CHARMm with VCharge charges, and the Poisson-Boltzmann Surface Area (PBSA) solvation model; and a semiempirical quantum mechanical Hamiltonian, PM6-DH+, coupled with the COSMO solvation model and a surface area term for nonpolar solvation free energy. Binding affinities based on the classical force field correlated strongly with the experiments with a correlation coefficient (R2) of 0.74. On the other hand, binding affinities based on the quantum mechanical energy model correlated poorly with experiments (R2 = 0.24), due largely to two major outliers. As we used extensive conformational search methods, these results point to possible inaccuracies in the PM6-DH+ energy model or the COSMO solvation model. Furthermore, the different binding free energy components, solute energy, solvation free energy, and configurational entropy showed significant deviations between the classical M2 and quantum M2 calculations. Comparison of different classical M2 free energy components to experiments show that the change in the total energy, i.e. the solute energy plus the solvation free energy, is the key driving force for binding, with a reasonable correlation to experiment (R2 = 0.56); however, accounting for configurational entropy further improves the correlation. PMID:24510191

  14. On the Presence of Affine Fibril and Fiber Kinematics in the Mitral Valve Anterior Leaflet

    PubMed Central

    Lee, Chung-Hao; Zhang, Will; Liao, Jun; Carruthers, Christopher A.; Sacks, Jacob I.; Sacks, Michael S.

    2015-01-01

    In this study, we evaluated the hypothesis that the constituent fibers follow an affine deformation kinematic model for planar collagenous tissues. Results from two experimental datasets were utilized, taken at two scales (nanometer and micrometer), using mitral valve anterior leaflet (MVAL) tissues as the representative tissue. We simulated MVAL collagen fiber network as an ensemble of undulated fibers under a generalized two-dimensional deformation state, by representing the collagen fibrils based on a planar sinusoidally shaped geometric model. The proposed approach accounted for collagen fibril amplitude, crimp period, and rotation with applied macroscopic tissue-level deformation. When compared to the small angle x-ray scattering measurements, the model fit the data well, with an r2 = 0.976. This important finding suggests that, at the homogenized tissue-level scale of ∼1 mm, the collagen fiber network in the MVAL deforms according to an affine kinematics model. Moreover, with respect to understanding its function, affine kinematics suggests that the constituent fibers are largely noninteracting and deform in accordance with the bulk tissue. It also suggests that the collagen fibrils are tightly bounded and deform as a single fiber-level unit. This greatly simplifies the modeling efforts at the tissue and organ levels, because affine kinematics allows a straightforward connection between the macroscopic and local fiber strains. It also suggests that the collagen and elastin fiber networks act independently of each other, with the collagen and elastin forming long fiber networks that allow for free rotations. Such freedom of rotation can greatly facilitate the observed high degree of mechanical anisotropy in the MVAL and other heart valves, which is essential to heart valve function. These apparently novel findings support modeling efforts directed toward improving our fundamental understanding of tissue biomechanics in healthy and diseased conditions

  15. High Affinity Binding of Indium and Ruthenium Ions by Gastrins

    PubMed Central

    Baldwin, Graham S.; George, Graham N.; Pushie, M. Jake

    2015-01-01

    The peptide hormone gastrin binds two ferric ions with high affinity, and iron binding is essential for the biological activity of non-amidated forms of the hormone. Since gastrins act as growth factors in gastrointestinal cancers, and as peptides labelled with Ga and In isotopes are increasingly used for cancer diagnosis, the ability of gastrins to bind other metal ions was investigated systematically by absorption spectroscopy. The coordination structures of the complexes were characterized by extended X-ray absorption fine structure (EXAFS) spectroscopy. Changes in the absorption of gastrin in the presence of increasing concentrations of Ga3+ were fitted by a 2 site model with dissociation constants (Kd) of 3.3 x 10−7 and 1.1 x 10−6 M. Although the absorption of gastrin did not change upon the addition of In3+ ions, the changes in absorbance on Fe3+ ion binding in the presence of indium ions were fitted by a 2 site model with Kd values for In3+ of 6.5 x 10−15 and 1.7 x 10−7 M. Similar results were obtained with Ru3+ ions, although the Kd values for Ru3+ of 2.6 x 10−13 and 1.2 x 10−5 M were slightly larger than observed for In3+. The structures determined by EXAFS all had metal:gastrin stoichiometries of 2:1 but, while the metal ions in the Fe, Ga and In complexes were bridged by a carboxylate and an oxygen with a metal-metal separation of 3.0–3.3 Å, the Ru complex clearly demonstrated a short range Ru—Ru separation, which was significantly shorter, at 2.4 Å, indicative of a metal-metal bond. We conclude that gastrin selectively binds two In3+ or Ru3+ ions, and that the affinity of the first site for In3+ or Ru3+ ions is higher than for ferric ions. Some of the metal ion-gastrin complexes may be useful for cancer diagnosis and therapy. PMID:26457677

  16. Extremely high negative electron affinity of diamond via magnesium adsorption

    NASA Astrophysics Data System (ADS)

    O'Donnell, K. M.; Edmonds, M. T.; Tadich, A.; Thomsen, L.; Stacey, A.; Schenk, A.; Pakes, C. I.; Ley, L.

    2015-07-01

    We report large negative electron affinity (NEA) on diamond (100) using magnesium adsorption on a previously oxygen-terminated surface. The measured NEA is up to (-2.01 ±0.05 ) eV, the largest reported negative electron affinity to date. Despite the expected close relationship between the surface chemistry of Mg and Li species on oxygen-terminated diamond, we observe differences in the adsorption properties between the two. Most importantly, a high-temperature annealing step is not required to activate the Mg-adsorbed surface to a state of negative electron affinity. Diamond surfaces prepared by this procedure continue to possess negative electron affinity after exposure to high temperatures, air, and even immersion in water.

  17. COMPARATIVE OXYGEN AFFINITY OF FISH AND MAMMALIAN MYOGLOBINS

    EPA Science Inventory

    Myoglobins from rat, coho salmon (Oncorhynchus kisutch), buffalo sculpin (Enophrys bison) hearts, and yellowfin tuna (Thunnus albacares) red skeletal muscle were partially purified and their O2 binding affinities determined. Commercially prepared sperm whale myoglobin was employe...

  18. Proton affinity of several basic non-standard amino acids

    NASA Astrophysics Data System (ADS)

    Rožman, Marko

    2012-08-01

    The structures and absolute proton affinities of several arginine (2-amino-3-guanidinopropionic acid, 2-amino-4-guanidinobutyric acid, homoarginine, citrulline and canavanine), histidine (1-methylhistidine and 3-methylhistidine) and lysine (2,3-diaminopropanoic acid, 2,4-diaminobutanoic acid, ornithine, 5-hydroxylysine, canaline and thialysine) homologues and analogues have been estimated using composite G3MP2B3 computational protocol. For a majority of here studied non-standard amino acids the gas-phase proton affinities were established for the first time, while for the others obtained values are used to improve the accuracy of the computational and experimental proton affinities reported previously. In addition, structures and proton affinities are discussed in order to rationalize their biological activity.

  19. Frontal affinity chromatography (FAC): theory and basic aspects.

    PubMed

    Kasai, Ken-ichi

    2014-01-01

    Frontal affinity chromatography (FAC) is a versatile analytical tool for determining specific interactions between biomolecules and is particularly useful in the field of glycobiology. This article presents its basic aspects, merits, and theory. PMID:25117240

  20. Antibody Affinity Maturation in Fishes—Our Current Understanding

    PubMed Central

    Magor, Brad G.

    2015-01-01

    It has long been believed that fish lack antibody affinity maturation, in part because they were thought to lack germinal centers. Recent research done on sharks and bony fishes indicates that these early vertebrates are able to affinity mature their antibodies. This article reviews the functionality of the fish homologue of the immunoglobulin (Ig) mutator enzyme activation-induced cytidine deaminase (AID). We also consider the protein and molecular evidence for Ig somatic hypermutation and antibody affinity maturation. In the context of recent evidence for a putative proto-germinal center in fishes we propose some possible reasons that observed affinity maturation in fishes often seems lacking and propose future work that might shed further light on this process in fishes. PMID:26264036

  1. Affinity regression predicts the recognition code of nucleic acid binding proteins

    PubMed Central

    Pelossof, Raphael; Singh, Irtisha; Yang, Julie L.; Weirauch, Matthew T.; Hughes, Timothy R.; Leslie, Christina S.

    2016-01-01

    Predicting the affinity profiles of nucleic acid-binding proteins directly from the protein sequence is a major unsolved problem. We present a statistical approach for learning the recognition code of a family of transcription factors (TFs) or RNA-binding proteins (RBPs) from high-throughput binding assays. Our method, called affinity regression, trains on protein binding microarray (PBM) or RNA compete experiments to learn an interaction model between proteins and nucleic acids, using only protein domain and probe sequences as inputs. By training on mouse homeodomain PBM profiles, our model correctly identifies residues that confer DNA-binding specificity and accurately predicts binding motifs for an independent set of divergent homeodomains. Similarly, learning from RNA compete profiles for diverse RBPs, our model can predict the binding affinities of held-out proteins and identify key RNA-binding residues. More broadly, we envision applying our method to model and predict biological interactions in any setting where there is a high-throughput ‘affinity’ readout. PMID:26571099

  2. Aptamer-modified magnetic beads in affinity separation of proteins.

    PubMed

    Zhu, Guohong; Walter, Johanna-Gabriela

    2015-01-01

    Aptamers are valuable alternative ligands for affinity separations. Here, we describe the aptamer-based affinity separation of His-tagged proteins using an aptamer directed against the His-tag. The immobilization of the aptamer to magnetic beads is described as well as the aptamer-based purification and proper methods for the characterization of the process. Moreover, indications for the transfer of the process to other aptamers are given. PMID:25749947

  3. Proton affinity of methyl nitrate - Less than proton affinity of nitric acid

    NASA Technical Reports Server (NTRS)

    Lee, Timothy J.; Rice, Julia E.

    1992-01-01

    Several state-of-the-art ab initio quantum mechanical methods were used to investigate the equilibrium structure, dipole moments, harmonic vibrational frequencies, and IR intensities of methyl nitrate, methanol, and several structures of protonated methyl nitrate, using the same theoretical methods as in an earlier study (Lee and Rice, 1992) of nitric acid. The ab initio results for methyl nitrate and methanol were found to be in good agreement with available experimental data. The proton affinity (PA) of methyl nitrate was calculated to be 176.9 +/-5 kcal/mol, in excellent agreement with the experimental value 176 kcal/mol obtained by Attina et al. (1987) and less than the PA value of nitric acid. An explanation of the discrepancy of the present results with those of an earlier study on protonated nitric acid is proposed.

  4. Ab initio determination of the proton affinities of small neutral and anionic molecules

    NASA Technical Reports Server (NTRS)

    DeFrees, D. J.; McLean, A. D.

    1986-01-01

    The proton affinity of a molecule in the gas phase is a fundamental measure of its basicity and is the factor controlling the course of many ion-molecule reactions. In this article, ab initio molecular orbital theory at the MP4/6-311 ++ G(3df, 3pd) level of theory is demonstrated to predict proton affinities (PA's) for small neutral and anionic bases to within 2 kcal mol-1. Furthermore, the errors are random, indicating that there are likely no systematic errors in either the experimental or theoretical PA's. Also, this level of theory is used to calibrate less sophisticated theoretical models which are suitable for larger molecules; the MP4/6-311 ++ G(2d, 2p) and MP2/6-311 ++ G(d, p) theoretical models should be particularly useful. A procedure for predicting the vibrational frequencies for anion is proposed and applied to CH3-, NH2-, OH-, and CN-.

  5. A novel method to estimate the affinity of HLA-A∗0201 restricted CTL epitope

    NASA Astrophysics Data System (ADS)

    Xu, Yun-sheng; Lin, Yong; Zhu, Bo; Lin, Zhi-hua

    2009-02-01

    A set of 70 peptides with affinity for the class I MHC HLA-A∗0201 molecule was subjected to quantitative structure-affinity relationship studies based on the SCORE function with good results ( r2 = 0.6982, RMS = 0.280). Then the 'leave-one-out' cross-validation (LOO-CV) and an outer test set including 18 outer samples were used to validate the QSAR model. The results of the LOO-CV were q2 = 0.6188, RMS = 0.315, and the results of outer test set were r2 = 0.5633, RMS = 0.2292. All these show that the QSAR model has good predictability. Statistical analysis showed that the hydrophobic and hydrogen bond interaction played a significant role in peptide-MHC molecule binding. The study also provided useful information for structure modification of CTL epitope, and laid theoretical base for molecular design of therapeutic vaccine.

  6. Application of a constrained regularization method to extraction of affinity distributions: proton and metal binding to humic substances.

    PubMed

    Orsetti, Silvia; Andrade, Estela María; Molina, Fernando V

    2009-08-15

    The binding of proton and metal cations to humic substances has been analyzed with a regularized fitting procedure (using the CONTIN software package) to extract conditional affinity distributions, valid at a given ionic strength, from binding (titration) curves. The procedure was previously tested with simulated titration curves using a simple bi-Gaussian model, the NICA-Donnan model, and the Stockholm humic model. Application to literature data for proton binding shows that in several cases the affinity distribution found is bimodal (carboxylic and phenolic sites) as usually assumed; however in other cases, specially for fulvic acids, a trimodal distribution is clearly discerned, with a smaller peak between the two noted above attributed to the presence of vicinal carboxylic groups. The analysis of metal binding curves has been performed in a few cases where the available data could be reliably processed, separating the proton affinity distribution and obtaining the conditional affinity spectra. For Cd(II) and Pb(II) a bimodal distribution is found, attributed in principle to mono- and bidentate binding, based on spectroscopic data. In the case of Cu(II), a more complex affinity distribution is found showing 3-4 peaks; this is consistent with spectroscopic studies, where different binding modes, up to tetradentate, have been observed. PMID:19477457

  7. Correlation between AcrB trimer association affinity and efflux activity.

    PubMed

    Ye, Cui; Wang, Zhaoshuai; Lu, Wei; Zhong, Meng; Chai, Qian; Wei, Yinan

    2014-06-17

    The majority of membrane proteins function as oligomers. However, it remains largely unclear how the oligomer stability of protein complexes correlates with their function. Understanding the relationship between oligomer stability and activity is essential to protein research and to virtually all cellular processes that depend on the function of protein complexes. Proteins make lasting or transient interactions as they perform their functions. Obligate oligomeric proteins exist and function exclusively at a specific oligomeric state. Although oligomerization is clearly critical for such proteins to function, a direct correlation between oligomer affinity and biological activity has not yet been reported. Here, we used an obligate trimeric membrane transporter protein, AcrB, as a model to investigate the correlation between its relative trimer affinity and efflux activity. AcrB is a component of the major multidrug efflux system in Escherichia coli. We created six AcrB constructs with mutations at the transmembrane intersubunit interface, and we determined their activities using both a drug susceptibility assay and an ethidium bromide accumulation assay. The relative trimer affinities of these mutants in detergent micelles were obtained using blue native polyacrylamide gel electrophoresis. A correlation between the relative trimer affinity and substrate efflux activity was observed, in which a threshold trimer stability was required to maintain efflux activity. The trimer affinity of the wild-type protein was approximately 3 kcal/mol more stable than the threshold value. Once the threshold was reached, an additional increase of stability in the range observed had no observable effect on protein activity. PMID:24854514

  8. Analysis and prediction of affinity of TAP binding peptides using cascade SVM.

    PubMed

    Bhasin, Manoj; Raghava, G P S

    2004-03-01

    The generation of cytotoxic T lymphocyte (CTL) epitopes from an antigenic sequence involves number of intracellular processes, including production of peptide fragments by proteasome and transport of peptides to endoplasmic reticulum through transporter associated with antigen processing (TAP). In this study, 409 peptides that bind to human TAP transporter with varying affinity were analyzed to explore the selectivity and specificity of TAP transporter. The abundance of each amino acid from P1 to P9 positions in high-, intermediate-, and low-affinity TAP binders were examined. The rules for predicting TAP binding regions in an antigenic sequence were derived from the above analysis. The quantitative matrix was generated on the basis of contribution of each position and residue in binding affinity. The correlation of r = 0.65 was obtained between experimentally determined and predicted binding affinity by using a quantitative matrix. Further a support vector machine (SVM)-based method has been developed to model the TAP binding affinity of peptides. The correlation (r = 0.80) was obtained between the predicted and experimental measured values by using sequence-based SVM. The reliability of prediction was further improved by cascade SVM that uses features of amino acids along with sequence. An extremely good correlation (r = 0.88) was obtained between measured and predicted values, when the cascade SVM-based method was evaluated through jackknife testing. A Web service, TAPPred (http://www.imtech.res.in/raghava/tappred/ or http://bioinformatics.uams.edu/mirror/tappred/), has been developed based on this approach. PMID:14978300

  9. Computational estimation of rainbow trout estrogen receptor binding affinities for environmental estrogens

    SciTech Connect

    Shyu, Conrad; Cavileer, Timothy D.; Nagler, James J.; Ytreberg, F. Marty

    2011-02-01

    Environmental estrogens have been the subject of intense research due to their documented detrimental effects on the health of fish and wildlife and their potential to negatively impact humans. A complete understanding of how these compounds affect health is complicated because environmental estrogens are a structurally heterogeneous group of compounds. In this work, computational molecular dynamics simulations were utilized to predict the binding affinity of different compounds using rainbow trout (Oncorhynchus mykiss) estrogen receptors (ERs) as a model. Specifically, this study presents a comparison of the binding affinity of the natural ligand estradiol-17{beta} to the four rainbow trout ER isoforms with that of three known environmental estrogens 17{alpha}-ethinylestradiol, bisphenol A, and raloxifene. Two additional compounds, atrazine and testosterone, that are known to be very weak or non-binders to ERs were tested. The binding affinity of these compounds to the human ER{alpha} subtype is also included for comparison. The results of this study suggest that, when compared to estradiol-17{beta}, bisphenol A binds less strongly to all four receptors, 17{alpha}-ethinylestradiol binds more strongly, and raloxifene has a high affinity for the {alpha} subtype only. The results also show that atrazine and testosterone are weak or non-binders to the ERs. All of the results are in excellent qualitative agreement with the known in vivo estrogenicity of these compounds in the rainbow trout and other fishes. Computational estimation of binding affinities could be a valuable tool for predicting the impact of environmental estrogens in fish and other animals.

  10. Enhancing IHE XDS for federated clinical affinity domain support.

    PubMed

    Dogac, Asuman; Laleci, Gokce B; Aden, Thomas; Eichelberg, Marco

    2007-03-01

    One of the key problems in healthcare informatics is the inability to share patient records across enterprises. To address this problem, an important industry initiative called "integrating the healthcare enterprise (IHE)" specified the "cross enterprise document sharing (XDS)" profile. In the IHE XDS, healthcare enterprises that agree to work together form a "clinical affinity domain" and store healthcare documents in an ebXML registry/repository architecture to facilitate their sharing. The affinity domains also agree on a common set of policies such as coding lists to be used to annotate clinical documents in the registry/repository and the common schemes for patient identification. However, since patients expect their records to follow them as they move from one clinical affinity domain to another, there is a need for affinity domains to be federated to enable information exchange. In this paper, we describe how IHE XDS can be enhanced to support federated clinical affinity domains. We demonstrate that federation of affinity domains are facilitated when ontologies, rather than coding term lists, are used to annotate clinical documents. Furthermore, we describe a patient identification protocol that eliminates the need to keep a master patient index file for the federation. PMID:17390991

  11. Flexible Linker Modulates Glycosaminoglycan Affinity of Decorin Binding Protein A.

    PubMed

    Morgan, Ashli; Sepuru, Krishna Mohan; Feng, Wei; Rajarathnam, Krishna; Wang, Xu

    2015-08-18

    Decorin binding protein A (DBPA) is a glycosaminoglycan (GAG)-binding adhesin found on the surface of the bacterium Borrelia burgdorferi (B. burgdorferi), the causative agent of Lyme disease. DBPA facilitates bacterial adherence to extracellular matrices of human tissues and is crucial during the early stage of the infection process. Interestingly, DBPA from different strains (B31, N40, and PBr) show significant differences in GAG affinities, but the structural basis for the differences is not clear. In this study, we show that GAG affinity of N40 DBPA is modulated in part by flexible segments that control access to the GAG binding site, such that shortening of the linker leads to higher GAG affinity when analyzed using ELISA, gel mobility shift assay, solution NMR, and isothermal titration calorimetry. Our observation that GAG affinity differences among different B. burgdorferi strains can be attributed to a flexible linker domain regulating access to the GAG-binding domain is novel. It also provides a rare example of how neutral amino acids and dynamic segments in GAG binding proteins can have a large influence on GAG affinity and provides insights into why the number of basic amino acids in the GAG-binding site may not be the only factor determining GAG affinity of proteins. PMID:26223367

  12. Experimental Immunization Based on Plasmodium Antigens Isolated by Antibody Affinity

    PubMed Central

    Kamali, Ali N.; Marín-García, Patricia; Azcárate, Isabel G.; Puyet, Antonio; Diez, Amalia; Bautista, José M.

    2015-01-01

    Vaccines blocking malaria parasites in the blood-stage diminish mortality and morbidity caused by the disease. Here, we isolated antigens from total parasite proteins by antibody affinity chromatography to test an immunization against lethal malaria infection in a murine model. We used the sera of malaria self-resistant ICR mice to lethal Plasmodium yoelii yoelii 17XL for purification of their IgGs which were subsequently employed to isolate blood-stage parasite antigens that were inoculated to immunize BALB/c mice. The presence of specific antibodies in vaccinated mice serum was studied by immunoblot analysis at different days after vaccination and showed an intensive immune response to a wide range of antigens with molecular weight ranging between 22 and 250 kDa. The humoral response allowed delay of the infection after the inoculation to high lethal doses of P. yoelii yoelii 17XL resulting in a partial protection against malaria disease, although final survival was managed in a low proportion of challenged mice. This approach shows the potential to prevent malaria disease with a set of antigens isolated from blood-stage parasites. PMID:26539558

  13. On boundary fusion and functional relations in the Baxterized affine Hecke algebra

    SciTech Connect

    Babichenko, A.; Regelskis, V.

    2014-04-15

    We construct boundary type operators satisfying fused reflection equation for arbitrary representations of the Baxterized affine Hecke algebra. These operators are analogues of the fused reflection matrices in solvable half-line spin chain models. We show that these operators lead to a family of commuting transfer matrices of Sklyanin type. We derive fusion type functional relations for these operators for two families of representations.

  14. Membrane modulates affinity for calcium ion to create an apparent cooperative binding response by annexin a5.

    PubMed

    Gauer, Jacob W; Knutson, Kristofer J; Jaworski, Samantha R; Rice, Anne M; Rannikko, Anika M; Lentz, Barry R; Hinderliter, Anne

    2013-06-01

    Isothermal titration calorimetry was used to characterize the binding of calcium ion (Ca²⁺) and phospholipid to the peripheral membrane-binding protein annexin a5. The phospholipid was a binary mixture of a neutral and an acidic phospholipid, specifically phosphatidylcholine and phosphatidylserine in the form of large unilamellar vesicles. To stringently define the mode of binding, a global fit of data collected in the presence and absence of membrane concentrations exceeding protein saturation was performed. A partition function defined the contribution of all heat-evolving or heat-absorbing binding states. We find that annexin a5 binds Ca²⁺ in solution according to a simple independent-site model (solution-state affinity). In the presence of phosphatidylserine-containing liposomes, binding of Ca²⁺ differentiates into two classes of sites, both of which have higher affinity compared with the solution-state affinity. As in the solution-state scenario, the sites within each class were described with an independent-site model. Transitioning from a solution state with lower Ca²⁺ affinity to a membrane-associated, higher Ca²⁺ affinity state, results in cooperative binding. We discuss how weak membrane association of annexin a5 prior to Ca²⁺ influx is the basis for the cooperative response of annexin a5 toward Ca²⁺, and the role of membrane organization in this response. PMID:23746516

  15. Using the concept of transient complex for affinity predictions in CAPRI rounds 20-27 and beyond.

    PubMed

    Qin, Sanbo; Zhou, Huan-Xiang

    2013-12-01

    Predictions of protein-protein binders and binding affinities have traditionally focused on features pertaining to the native complexes. In developing a computational method for predicting protein-protein association rate constants, we introduced the concept of transient complex after mapping the interaction energy surface. The transient complex is located at the outer boundary of the bound-state energy well, having near-native separation and relative orientation between the subunits but not yet formed most of the short-range native interactions. We found that the width of the binding funnel and the electrostatic interaction energy of the transient complex are among the features predictive of binders and binding affinities. These ideas were very promising for the five affinity-related targets (T43-45, 55, and 56) of CAPRI rounds 20-27. For T43, we ranked the single crystallographic complex as number 1 and were one of only two groups that clearly identified that complex as a true binder; for T44, we ranked the only design with measurable binding affinity as number 4. For the nine docking targets, continuing on our success in previous CAPRI rounds, we produced 10 medium-quality models for T47 and acceptable models for T48 and T49. We conclude that the interaction energy landscape and the transient complex in particular will complement existing features in leading to better prediction of binding affinities. PMID:23873496

  16. Affinities of methylphenidate derivatives for dopamine, norepinephrine and serotonin transporters.

    PubMed

    Gatley, S J; Pan, D; Chen, R; Chaturvedi, G; Ding, Y S

    1996-01-01

    We have synthesized several derivative of dl-threo-methylphenidate (Ritalin) bearing substituents on the phenyl ring. IC50 values for binding these compounds to rat brain monoamine transporters were assessed using [3H]WIN 35,428 (striatal membranes, dopamine transporters, DAT), [3H]nisoxetine (frontal cortex membranes, norepinephrine transporters, NET) and [3H]paroxetine (brain stem membranes, 5HT transporters, 5HTT). Affinities (1/Ki) decreased in the order: DAT > NET > 5HTT. Substitution at the para position of dl-threo-methylphenidate generally led to retained or increased affinity for the dopamine transporter (bromo > iodo > methoxy > hydroxy). Substitution at the meta position also increased affinity for the DAT (m-bromo > methylphenidate; m-iodo-p-hydroxy > p-hydroxy). Substitution at the ortho position with bromine considerably decreased affinity. Similar IC50 values for binding of o-bromomethylphenidate to the dopamine transporter were measured at 0, 22 and 37 degrees. N-Methylation of the piperidine ring of methylphenidate also considerably reduced affinity. The dl-erythro isomer of o-bromomethylphenidate did not bind to the DAT (IC50 > 50,000 nM). Affinities at the dopamine and norepinephrine transporters for substituted methylphenidate derivatives were well correlated (r2=0.90). Abilities of several methylphenidate derivatives to inhibit [3H]dopamine uptake in striatal synaptosomes corresponded well with inhibition of [3H]WIN 35, 428 binding. None of the compounds examined exhibited significant affinity to dopamine D1 or D2 receptors (IC50 > 500 or 5,000 nM, respectively), as assessed by inhibition of binding of [3H]SCH 23390 or [123I]epidepride, respectively, to striatal membranes. PMID:8786705

  17. Hortonian scaling and effect of cutoffs on statistics of self-affine river networks

    NASA Astrophysics Data System (ADS)

    Neuman, Shlomo P.

    2009-12-01

    The planar structure of river networks exhibits fractal properties. In particular, available data indicate a tendency for drainage areas A to be distributed according to a power law; their boundaries and main channels to form self-affine curves; the characteristic lengths of drainage areas to be independently self-affine in the one-dimensional space Z of total channel length, rendering the network elongated (and anisotropic); the inverse network density A/Z to be either constant or self-affine in Z, depending on whether or not the network fills the two-dimensional space of A; and (as found in a recent study) areas of given size, vegetative cover, and mean steady state soil moisture, weighted by their flow distance from the basin outlet, to be self-affine in the one-dimensional space of this distance. Various theoretical and semiempirical relationships have been proposed among exponents defining some of these and other scale dependencies. Expressions have been proposed for ensemble moments of A and some length measures associated with finite size river basins. We present a new model that views any self-affine basin property, Y(X), as belonging to an infinite hierarchy of mutually uncorrelated, statistically homogeneous random functions defined on elementary subbasins, each of which is characterized by a unique integral (spatial correlation) scale λ. We cite a mathematically rigorous hydrologic rationale for our model and use it in conjunction with published scaling relations to obtain the probability density function of λ; to derive analytical expressions for ensemble analogs of Horton's scaling laws; to deduce from them that streams of any Horton-Strahler order ω are associated with integral scales λω ≤ λ ≤ λω+1, where the ratio λω+1/λω is a constant independent of ω; to develop analytical expressions relating statistical moments of Y(X) to arbitrary lower and upper cutoff scales that may (but need not) be taken to represent data support and

  18. Binding affinity prediction of novel estrogen receptor ligands using receptor-based 3-D QSAR methods.

    PubMed

    Sippl, Wolfgang

    2002-12-01

    We have recently reported the development of a 3-D QSAR model for estrogen receptor ligands showing a significant correlation between calculated molecular interaction fields and experimentally measured binding affinity. The ligand alignment obtained from docking simulations was taken as basis for a comparative field analysis applying the GRID/GOLPE program. Using the interaction field derived with a water probe and applying the smart region definition (SRD) variable selection procedure, a significant and robust model was obtained (q(2)(LOO)=0.921, SDEP=0.345). To further analyze the robustness and the predictivity of the established model several recently developed estrogen receptor ligands were selected as external test set. An excellent agreement between predicted and experimental binding data was obtained indicated by an external SDEP of 0.531. Two other traditionally used prediction techniques were applied in order to check the performance of the receptor-based 3-D QSAR procedure. The interaction energies calculated on the basis of receptor-ligand complexes were correlated with experimentally observed affinities. Also ligand-based 3-D QSAR models were generated using program FlexS. The interaction energy-based model, as well as the ligand-based 3-D QSAR models yielded models with lower predictivity. The comparison with the interaction energy-based model and with the ligand-based 3-D QSAR models, respectively, indicates that the combination of receptor-based and 3-D QSAR methods is able to improve the quality of prediction. PMID:12413831

  19. Characterization of a dockerin-based affinity tag: application for purification of a broad variety of target proteins.

    PubMed

    Demishtein, Alik; Karpol, Alon; Barak, Yoav; Lamed, Raphael; Bayer, Edward A

    2010-01-01

    Cellulose, a major component of plant matter, is degraded by a cell surface multiprotein complex called the cellulosome produced by several anaerobic bacteria. This complex coordinates the assembly of different glycoside hydrolases, via a high-affinity Ca(2+)-dependent interaction between the enzyme-borne dockerin and the scaffoldin-borne cohesin modules. In this study, we characterized a new protein affinity tag, ΔDoc, a truncated version (48 residues) of the Clostridium thermocellum Cel48S dockerin. The truncated dockerin tag has a binding affinity (K(A)) of 7.7 × 10(8)M(-1), calculated by a competitive enzyme-linked assay system. In order to examine whether the tag can be used for general application in affinity chromatography, it was fused to a range of target proteins, including Aequorea victoria green fluorescent protein (GFP), C. thermocellum β-glucosidase, Escherichia coli thioesterase/protease I (TEP1), and the antibody-binding ZZ-domain from Staphylococcus aureus protein A. The results of this study significantly extend initial studies performed using the Geobacillus stearothermophilus xylanase T-6 as a model system. In addition, the enzymatic activity of a C. thermocellum β-glucosidase, purified using this approach, was tested and found to be similar to that of a β-glucosidase preparation (without the ΔDoc tag) purified using the standard His-tag. The truncated dockerin derivative functioned as an effective affinity tag through specific interaction with a cognate cohesin, and highly purified target proteins were obtained in a single step directly from crude cell extracts. The relatively inexpensive beaded cellulose-based affinity column was reusable and maintained high capacity after each cycle. This study demonstrates that deletion into the first Ca(2+)-binding loop of the dockerin module results in an efficient and robust affinity tag that can be generally applied for protein purification. PMID:21038354

  20. Robust Affinity Standards for Cu(I) Biochemistry

    PubMed Central

    Bagchi, Pritha; Morgan, M. Thomas; Bacsa, John; Fahrni, Christoph J.

    2014-01-01

    The measurement of reliable Cu(I) protein binding affinities requires competing reference ligands with similar binding strengths; however, the literature on such reference ligands is not only sparse but often conflicting. To address this deficiency, we have created and characterized a series of water-soluble monovalent copper ligands, MCL-1, MCL-2, and MCL-3, that form well-defined, air-stable, and colorless complexes with Cu(I) in aqueous solution. Concluding from X-ray structural data, electrochemical measurements, and an extensive network of equilibrium titrations, all three ligands form discrete Cu(I) complexes with 1:1 stoichiometry and are capable of buffering Cu(I) concentrations between 10−10 and 10−17 M. As most Cu(I) protein affinities have been obtained from competition experiments with bathocuproine disulfonate (BCS) or 2,2′-bicinchoninic acid (BCA), we further calibrated their Cu(I) stability constants against the MCL-series. To demonstrate the application of these reagents, we determined the Cu(I) binding affinity of CusF (logK = 14.3±0.1), a periplasmic metalloprotein required for the detoxification of elevated copper levels in E. coli. Altogether, this interconnected set of affinity standards establishes a reliable foundation that will facilitate the precise determination of Cu(I) binding affinities of proteins and small molecule ligands. PMID:24298878

  1. Coenzyme-like ligands for affinity isolation of cholesterol oxidase.

    PubMed

    Xin, Yu; Lu, Liushen; Wang, Qing; Zhang, Ling; Tong, Yanjun; Wang, Wu

    2016-05-15

    Two coenzyme-like chemical ligands were designed and synthesized for affinity isolation of cholesterol oxidase (COD). To simulate the structure of natural coenzyme of COD (flavin adenine dinucleotide (FAD)), on Sepharose beads, 5-aminouracil, cyanuric chloride and 1, 4-butanediamine were composed and then modified. The COD gene from Brevibacterium sp. (DQ345780) was expressed in Escherichia coli BL21 (DE3), and then the sorbents were applied to adsorption analysis with the pure enzyme. Subsequently, the captured enzyme was applied to SDS-PAGE and activity analysis. As calculated, the theoretical maximum adsorption (Qmax) of the two affinity sorbents (RL-1 and RL-2) were ∼83.5 and 46.3mg/g wet gel; and the desorption constant Kd of the two sorbents were ∼6.02×10(-4) and 1.19×10(-4)μM. The proteins after cell lysis were applied to affinity isolation, and then after one step of affinity binding on the two sorbents, the protein recoveries of RL-1 and RL-2 were 9.2% and 9.7%; the bioactivity recoveries were 92.7% and 91.3%, respectively. SDS-PAGE analysis revealed that the purities of COD isolated with the two affinity sorbents were approximately 95%. PMID:26856529

  2. Preparation of a novel Zr(4+)-immobilized metal affinity membrane for selective adsorption of phosphoprotein.

    PubMed

    He, Maofang; Wang, Chaozhan; Wei, Yinmao

    2016-09-01

    In this study, a novel phosphate-Zr(4+) immobilized metal affinity membrane (IMAM) was prepared based on the surface initiated-atom transfer radical polymerization technique for the selective adsorption of phosphoprotein. The adsorption capacity and selectivity of the phosphate-Zr(4+) IMAM were evaluated by using the mixture of standard phosphoproteins (β-casein, ovalbumin) and nonphosphoproteins (bovine serum albumin and lysozyme) as model samples. The adsorption isotherms and competitive adsorption results demonstrated that the phosphate-Zr(4+) IMAM had higher binding capacity and selectivity for phosphoproteins over nonphosphoproteins. Moreover, the phosphate-Zr(4+) IMAM exhibited good re-usability and re-productivity. Finally, the phosphate-Zr(4+) IMAM was applied to separate phosphoprotein from real samples with high purity. Therefore, the as-prepared phosphate-Zr(4+) IMAM could be a promising affinity material for the efficient enrichment of phosphoprotein from complex bio-samples. PMID:27433983

  3. Particle displacements in the elastic deformation of amorphous materials: Local fluctuations vs. non-affine field

    NASA Astrophysics Data System (ADS)

    Goldenberg, C.; Tanguy, A.; Barrat, J.-L.

    2007-10-01

    We study the local disorder in the deformation of amorphous materials by decomposing the particle displacements into a continuous, inhomogeneous field and the corresponding fluctuations. We compare these fields to the commonly used non-affine displacements in an elastically deformed 2D Lennard-Jones glass. Unlike the non-affine field, the fluctuations are very localized, and exhibit a much smaller (and system size independent) correlation length, on the order of a particle diameter, supporting the applicability of the notion of local "defects" to such materials. We propose a scalar "noise" field to characterize the fluctuations, as an additional field for extended continuum models, e.g., to describe the localized irreversible events observed during plastic deformation.

  4. Structure-based Design of Peptides with High Affinity and Specificity to HER2 Positive Tumors

    PubMed Central

    Geng, Lingling; Wang, Zihua; Yang, Xiaoliang; Li, Dan; Lian, Wenxi; Xiang, Zhichu; Wang, Weizhi; Bu, Xiangli; Lai, Wenjia; Hu, Zhiyuan; Fang, Qiaojun

    2015-01-01

    To identify peptides with high affinity and specificity against human epidermal growth factor receptor 2 (HER2), a series of peptides were designed based on the structure of HER2 and its Z(HER2:342) affibody. By using a combination protocol of molecular dynamics modeling, MM/GBSA binding free energy calculations, and binding free energy decomposition analysis, two novel peptides with 27 residues, pep27 and pep27-24M, were successfully obtained. Immunocytochemistry and flow cytometry analysis verified that both peptides can specifically bind to the extracellular domain of HER2 protein at cellular level. The Surface Plasmon Resonance imaging (SPRi) analysis showed that dissociation constants (KD) of these two peptides were around 300 nmol/L. Furthermore, fluorescence imaging of peptides against nude mice xenografted with SKBR3 cells indicated that both peptides have strong affinity and high specificity to HER2 positive tumors. PMID:26284145

  5. Low-complexity piecewise-affine virtual sensors: theory and design

    NASA Astrophysics Data System (ADS)

    Rubagotti, Matteo; Poggi, Tomaso; Oliveri, Alberto; Pascucci, Carlo Alberto; Bemporad, Alberto; Storace, Marco

    2014-03-01

    This paper is focused on the theoretical development and the hardware implementation of low-complexity piecewise-affine direct virtual sensors for the estimation of unmeasured variables of interest of nonlinear systems. The direct virtual sensor is designed directly from measured inputs and outputs of the system and does not require a dynamical model. The proposed approach allows one to design estimators which mitigate the effect of the so-called 'curse of dimensionality' of simplicial piecewise-affine functions, and can be therefore applied to relatively high-order systems, enjoying convergence and optimality properties. An automatic toolchain is also presented to generate the VHDL code describing the digital circuit implementing the virtual sensor, starting from the set of measured input and output data. The proposed methodology is applied to generate an FPGA implementation of the virtual sensor for the estimation of vehicle lateral velocity, using a hardware-in-the-loop setting.

  6. Structure-based Design of Peptides with High Affinity and Specificity to HER2 Positive Tumors.

    PubMed

    Geng, Lingling; Wang, Zihua; Yang, Xiaoliang; Li, Dan; Lian, Wenxi; Xiang, Zhichu; Wang, Weizhi; Bu, Xiangli; Lai, Wenjia; Hu, Zhiyuan; Fang, Qiaojun

    2015-01-01

    To identify peptides with high affinity and specificity against human epidermal growth factor receptor 2 (HER2), a series of peptides were designed based on the structure of HER2 and its Z(HER2:342) affibody. By using a combination protocol of molecular dynamics modeling, MM/GBSA binding free energy calculations, and binding free energy decomposition analysis, two novel peptides with 27 residues, pep27 and pep27-24M, were successfully obtained. Immunocytochemistry and flow cytometry analysis verified that both peptides can specifically bind to the extracellular domain of HER2 protein at cellular level. The Surface Plasmon Resonance imaging (SPRi) analysis showed that dissociation constants (K D) of these two peptides were around 300 nmol/L. Furthermore, fluorescence imaging of peptides against nude mice xenografted with SKBR3 cells indicated that both peptides have strong affinity and high specificity to HER2 positive tumors. PMID:26284145

  7. Optimizing the affinity and specificity of ligand binding with the inclusion of solvation effect.

    PubMed

    Yan, Zhiqiang; Wang, Jin

    2015-09-01

    Solvation effect is an important factor for protein-ligand binding in aqueous water. Previous scoring function of protein-ligand interactions rarely incorporates the solvation model into the quantification of protein-ligand interactions, mainly due to the immense computational cost, especially in the structure-based virtual screening, and nontransferable application of independently optimized atomic solvation parameters. In order to overcome these barriers, we effectively combine knowledge-based atom-pair potentials and the atomic solvation energy of charge-independent implicit solvent model in the optimization of binding affinity and specificity. The resulting scoring functions with optimized atomic solvation parameters is named as specificity and affinity with solvation effect (SPA-SE). The performance of SPA-SE is evaluated and compared to 20 other scoring functions, as well as SPA. The comparative results show that SPA-SE outperforms all other scoring functions in binding affinity prediction and "native" pose identification. Our optimization validates that solvation effect is an important regulator to the stability and specificity of protein-ligand binding. The development strategy of SPA-SE sets an example for other scoring function to account for the solvation effect in biomolecular recognitions. PMID:26111900

  8. Predicting Adsorption Affinities of Small Molecules on Carbon Nanotubes Using Molecular Dynamics Simulation.

    PubMed

    Comer, Jeffrey; Chen, Ran; Poblete, Horacio; Vergara-Jaque, Ariela; Riviere, Jim E

    2015-12-22

    Computational techniques have the potential to accelerate the design and optimization of nanomaterials for applications such as drug delivery and contaminant removal; however, the success of such techniques requires reliable models of nanomaterial surfaces as well as accurate descriptions of their interactions with relevant solutes. In the present work, we evaluate the ability of selected models of naked and hydroxylated carbon nanotubes to predict adsorption equilibrium constants for about 30 small aromatic compounds with a variety of functional groups. The equilibrium constants determined using molecular dynamics coupled with free-energy calculation techniques are directly compared to those derived from experimental measurements. The calculations are highly predictive of the relative adsorption affinities of the compounds, with excellent correlation (r ≥ 0.9) between calculated and measured values of the logarithm of the adsorption equilibrium constant. Moreover, the agreement in absolute terms is also reasonable, with average errors of less than one decade. We also explore possible effects of surface loading, although we demonstrate that they are negligible for the experimental conditions considered. Given the degree of reliability demonstrated, we move on to employing the in silico techniques in the design of nanomaterials, using the optimization of adsorption affinity for the herbacide atrazine as an example. Our simulations suggest that, compared to other modifications of graphenic carbon, polyvinylpyrrolidone conjugation gives the highest affinity for atrazine-substantially greater than that of graphenic carbon alone-and may be useful as a nanomaterial for delivery or sequestration of atrazine. PMID:26506132

  9. SnAvi--a new tandem tag for high-affinity protein-complex purification.

    PubMed

    Schäffer, Ursula; Schlosser, Andreas; Müller, Kristian M; Schäfer, Angelika; Katava, Nenad; Baumeister, Ralf; Schulze, Ekkehard

    2010-04-01

    Systematic tandem-affinity-purification (TAP) of protein complexes was tremendously successful in yeast and has changed the general concept of how we understand protein function in eukaryotic cells. The transfer of this method to other model organisms has been difficult and may require specific adaptations. We were especially interested to establish a cell-type-specific TAP system for Caenorhabditis elegans, a model animal well suited to high-throughput analysis, proteomics and systems biology. By combining the high-affinity interaction between in vivo biotinylated target-proteins and streptavidin with the usage of a newly identified epitope of the publicly shared SB1 monoclonal antibody we created a novel in vivo fluorescent tag, the SnAvi-Tag. We show the versatile application of the SnAvi-Tag in Escherichia coli, vertebrate cells and in C. elegans for tandem affinity purification of protein complexes, western blotting and also for the in vivo sub-cellular localization of labelled proteins. PMID:20047968

  10. Fluorescent measurement of affinity binding between thrombin and its aptamers using on-chip affinity monoliths.

    PubMed

    Gao, Changlu; Sun, Xiuhua; Woolley, Adam T

    2013-05-24

    A microfluidic chip with integrated 2mm long monoliths incorporated with poly(ethylene glycol) (PEG) groups was developed for thrombin-aptamer interaction study. The non-G quartet forming oligonucleotide coated monoliths was compared to a 15 mer thrombin-binding aptamer, in which affinity binding and elution processes were real-time monitored fluorescently. The results showed that the fluorescence intensity of aptamer stationary phase is approximately 10 times higher than that of the control column, which is probably due to the successful suppression of nonspecific adsorption between thrombin and aptamers/monoliths by using PEG-monolith. The experiment was repeated using human serum albumin (HSA) and green fluorescence protein (GFP) as interferences, it was double confirmed that thrombin was selectively retained by PEG-monolith. An elution efficiency of 75% was achieved with an elute of 200mM acetic acid and 2M NaCI, and the eluted thrombin was successfully separated in an ionic buffer system of 20mM NaHCO3 (pH 9.5) with 3% PEG. The hydrophilic and antifouling properties of PEG-monolith greatly decrease nonspecific adsorption and enhance detection sensitivity, which provided an alternative method to perform on-chip fluorescent measurement of bioaffinity binding. PMID:23587316

  11. Craniodental affinities of Southeast Asia's "negritos" and the concordance with their genetic affinities.

    PubMed

    Bulbeck, David

    2013-01-01

    Genetic research into Southeast Asia's "negritos" has revealed their deep-rooted ancestry, with time depth comparable to that of Southwest Pacific populations. This finding is often interpreted as evidence that negritos, in contrast to other Southeast Asians, can trace much of their ancestry directly back to the early dispersal of Homo sapiens in the order of 70 kya from Africa to Pleistocene New Guinea and Australia. One view on negritos is to lump them and Southwest Pacific peoples into an "Australoid" race whose geographic distribution had included Southeast Asia prior to the Neolithic incursion of "Mongoloid" farmers. Studies into Semang osteology have revealed some hints of Southwest Pacific affinities in cranial shape, dental morphology, and dental metrical "shape." On the other hand, the Andamanese have been shown to resemble Africans in their craniometrics and South Asians in their dental morphology, while Philippine negritos resemble Mongoloid Southeast Asians in these respects and also in their dental metrics. This study expands the scope of negrito cranial comparisons by including Melayu Malays and additional coverage of South Asians. It highlights the distinction between the Mongoloid-like Philippine negritos and the Andamanese and Semang (and Senoi of Malaya) with their non-Mongoloid associations. It proposes that the early/mid-Holocene dispersal of the B4a1a mitochondrial DNA clade across Borneo, the Philippines, and Taiwan may be important for understanding the distinction between Philippine and other negritos. PMID:24297222

  12. Analysis of free drug fractions in serum by ultrafast affinity extraction and two-dimensional affinity chromatography using α1-acid glycoprotein microcolumns.

    PubMed

    Bi, Cong; Zheng, Xiwei; Hage, David S

    2016-02-01

    In the circulatory system, many drugs are reversibly bound to serum proteins such as human serum albumin (HSA) and alpha1-acid glycoprotein (AGP), resulting in both free and protein-bound fractions for these drugs. This report examined the use of microcolumns containing immobilized AGP for the measurement of free drug fractions by ultrafast affinity extraction and a two-dimensional affinity system. Several drugs known to bind AGP were used as models to develop and evaluate this approach. Factors considered during the creation of this method included the retention of the drugs on the microcolumns, the injection flow rate, the microcolumn size, and the times at which a second AGP column was placed on-line with the microcolumn. The final system had residence times of only 110-830ms during sample passage through the AGP microcolumns and allowed free drug fractions to be determined within 10-20min when using only 3-10μL of sample per injection. This method was used to measure the free fractions of the model drugs at typical therapeutic levels in serum, giving good agreement with the results obtained by ultrafiltration. This approach was also used to estimate the binding constants for each drug with AGP in serum, even for drugs that had significant interactions with both AGP and HSA in such samples. These results indicated that AGP microcolumns could be used with ultrafast affinity extraction to measure free drug fractions in a label-free manner and to study the binding of drugs with AGP in complex samples such as serum. PMID:26797422

  13. Enhancing Community Detection By Affinity-based Edge Weighting Scheme

    SciTech Connect

    Yoo, Andy; Sanders, Geoffrey; Henson, Van; Vassilevski, Panayot

    2015-10-05

    Community detection refers to an important graph analytics problem of finding a set of densely-connected subgraphs in a graph and has gained a great deal of interest recently. The performance of current community detection algorithms is limited by an inherent constraint of unweighted graphs that offer very little information on their internal community structures. In this paper, we propose a new scheme to address this issue that weights the edges in a given graph based on recently proposed vertex affinity. The vertex affinity quantifies the proximity between two vertices in terms of their clustering strength, and therefore, it is ideal for graph analytics applications such as community detection. We also demonstrate that the affinity-based edge weighting scheme can improve the performance of community detection algorithms significantly.

  14. Specific capture of uranyl protein targets by metal affinity chromatography.

    PubMed

    Basset, Christian; Dedieu, Alain; Guérin, Philippe; Quéméneur, Eric; Meyer, Daniel; Vidaud, Claude

    2008-03-28

    To improve general understanding of biochemical mechanisms in the field of uranium toxicology, the identification of protein targets needs to be intensified. Immobilized metal affinity chromatography (IMAC) has been widely developed as a powerful tool for capturing metal binding proteins from biological extracts. However uranyl cations (UO2(2+)) have particular physico-chemical characteristics which prevent them from being immobilized on classical metal chelating supports. We report here on the first development of an immobilized uranyl affinity chromatography method, based on the cation-exchange properties of aminophosphonate groups for uranyl binding. The cation distribution coefficient and loading capacity on the support were determined. Then the stability of the uranyl-bonded phase under our chromatographic conditions was optimized to promote affinity mechanisms. The successful enrichment of uranyl binding proteins from human serum was then proven using proteomic and mass spectral analysis. PMID:18308325

  15. Recent advances in affinity capillary electrophoresis for binding studies.

    PubMed

    Albishri, Hassan M; El Deeb, Sami; AlGarabli, Noura; AlAstal, Raghda; Alhazmi, Hassan A; Nachbar, Markus; El-Hady, Deia Abd; Wätzig, Hermann

    2014-01-01

    The present review covers recent advances and important applications of affinity capillary electrophoresis (ACE). It provides an overview about various ACE types, including ACE-MS, the multiple injection mode, the use of microchips and field-amplified sample injection-ACE. The most common scenarios of the studied affinity interactions are protein-drug, protein-metal ion, protein-protein, protein-DNA, protein-carbohydrate, carbohydrate-drug, peptide-peptide, DNA-drug and antigen-antibody. Approaches for the improvements of ACE in term of precision, rinsing protocols and sensitivity are discussed. The combined use of computer simulation programs to support data evaluation is presented. In conclusion, the performance of ACE is compared with other techniques such as equilibrium dialysis, parallel artificial membrane permeability assay, high-performance affinity chromatography as well as surface plasmon resonance, ultraviolet, circular dichroism, nuclear magnetic resonance, Fourier transform infrared, fluorescence, MS and isothermal titration calorimetry. PMID:25534793

  16. On the electron affinity of the oxygen atom

    NASA Technical Reports Server (NTRS)

    Bauschlicher, C. W., Jr.; Langhoff, S. R.; Partridge, H.; Taylor, P. R.

    1986-01-01

    The electron affinity (EA) of oxygen is computed to be 1.287 eV, using 2p electron full configuration-interaction (CI) wave functions expanded in a 6s5p3d2f Slater-type orbital basis. The best complete active space self-consistent field - multireference CI (CASSCF-MRCI) result including only 2p correlation is 1.263 eV. However, inclusion of 2s intrashell and 2s2p intershell correlation increases the computed EA to 1.290 at the CASSCF-MRCI level. At the full CI basis set limit, the 2s contribution to the electron affinity is estimated to be as large as 0.1 eV. This study clearly establishes the synergistic effect between the higher excitations and basis set completeness on the electron affinity when the 2s electrons are correlated.

  17. Gradient representations and affine structures in AEn

    NASA Astrophysics Data System (ADS)

    Kleinschmidt, Axel; Nicolai, Hermann

    2005-11-01

    We study the indefinite Kac Moody algebras AEn, arising in the reduction of Einstein's theory from (n + 1) spacetime dimensions to one (time) dimension, and their distinguished maximal regular subalgebras A_{n-1}\\equiv\\mathfrak{sl}_n and A(1)n-2. The interplay between these two subalgebras is used, for n = 3, to determine the commutation relations of the 'gradient generators' within AE3. The low-level truncation of the geodesic σ-model over the coset space AEn/K(AEn) is shown to map to a suitably truncated version of the SL(n)/SO(n) nonlinear σ-model resulting from the reduction Einstein's equations in (n + 1) dimensions to (1 + 1) dimensions. A further truncation to diagonal solutions can be exploited to define a one-to-one correspondence between such solutions, and null geodesic trajectories on the infinite-dimensional coset space {\\mathfrak{H}}/K({\\mathfrak{H}}) , where {\\mathfrak{H}} is the (extended) Heisenberg group, and K({\\mathfrak{H}}) its maximal compact subgroup. We clarify the relation between {\\mathfrak{H}} and the corresponding subgroup of the Geroch group.

  18. Surface plasmon resonance spectroscopy-based high-throughput screening of ligands for use in affinity and displacement chromatography.

    PubMed

    Vutukuru, Srinavya; Kane, Ravi S

    2008-10-21

    We describe an approach that uses surface plasmon resonance (SPR) spectroscopy and self-assembled monolayers (SAMs) for the high-throughput screening of ligands for use in displacement and affinity chromatographic processes. We identified a set of commercially available organic amines and allowed them to react with SAMs presenting interchain carboxylic anhydride groups; the resulting surfaces presented ligands of interest in a background of carboxylic acid groups. We used SPR spectroscopy to determine the extent of adsorption of two model proteinslysozyme and cytochrome conto these "multimodal" surfaces and to select promising "affinity" ligands for further characterization. The attachment of selected ligands to UltraLink Biosupport resulted in beads with a significantly greater affinity for lysozyme than for cytochrome c that would be suitable for use in affinity chromatographic processes. Furthermore, we also used the screens to design "affinity displacers"small molecules that selectively retain lysozyme on chromatographic resins, while displacing cytochrome c. The combination of SPR spectroscopy and SAMs represents a powerful technique for identifying novel ligands that enable the purification of complex protein mixtures. PMID:18788766

  19. Bilirubin removal from human plasma by Cibacron Blue F3GA using immobilized microporous affinity membranous capillary method.

    PubMed

    Zhang, Lei; Jin, Gu

    2005-07-01

    A novel affinity sorbent system for direct bilirubin removal from human plasma was developed. These new adsorbents comprise Cibacron Blue F3GA as the specific ligand, and microporous membranous poly(tetrafluoroethylene) capillary (modified by coating with a hydrophilic layer of poly(vinyl alcohol) after activation) as the carrier matrix. The affinity adsorbents carrying 126.5 micromol Cibacron Blue F3GA/g polymer was then used to remove bilirubin in a flow-injection system. Non-specific adsorption on the poly(vinyl alcohol) coated capillary remains low, and higher affinity adsorption capacity, of up to 76.2 mg/g polymer was obtained after dye immobilization. The bilirubin adsorption capacity of the affinity capillary decreased with increase in the recirculation rate of plasma. The adsorption capacity increased with increase the temperature while decreased with increase the ionic strength. The maximum adsorption was only observed in neutral solution (pH 6-7). The adsorption isotherm fitted the Langmuir model well. These new adsorbents have higher velocity of mass transfer, better adsorption capacity, less fouling, longer service life and good reusability. The results of blood tests suggested the dye affinity capillary has good blood compatibility. PMID:15894520

  20. Ferromagnetic levan composite: an affinity matrix to purify lectin.

    PubMed

    Angeli, Renata; da Paz, Nathalia V N; Maciel, Jackeline C; Araújo, Flávia F B; Paiva, Patrícia M G; Calazans, Glícia M T; Valente, Ana Paula; Almeida, Fábio C L; Coelho, Luana C B B; Carvalho, Luiz B; Silva, Maria da Paz C; dos Santos Correia, Maria Tereza

    2009-01-01

    A simple and inexpensive procedure used magnetite and levan to synthesize a composite recovered by a magnetic field. Lectins from Canavalia ensiformis (Con A) and Cratylia mollis (Cramoll 1 and Cramoll 1, 4) did bind specifically to composite. The magnetic property of derivative favored washing out contaminating proteins and recovery of pure lectins with glucose elution. Cramoll 1 was purified by this affinity binding procedure in two steps instead of a previous three-step protocol with ammonium sulfate fractionation, affinity chromatography on Sephadex G-75, and ion exchange chromatography through a CM-cellulose column. PMID:19547713

  1. Ferromagnetic Levan Composite: An Affinity Matrix to Purify Lectin

    PubMed Central

    Angeli, Renata; da Paz, Nathalia V. N.; Maciel, Jackeline C.; Araújo, Flávia F. B.; Paiva, Patrícia M. G.; Calazans, Glícia M. T.; Valente, Ana Paula; Almeida, Fábio C. L.; Coelho, Luana C. B. B.; Carvalho, Luiz B.; Silva, Maria da Paz C.; dos Santos Correia, Maria Tereza

    2009-01-01

    A simple and inexpensive procedure used magnetite and levan to synthesize a composite recovered by a magnetic field. Lectins from Canavalia ensiformis (Con A) and Cratylia mollis (Cramoll 1 and Cramoll 1, 4) did bind specifically to composite. The magnetic property of derivative favored washing out contaminating proteins and recovery of pure lectins with glucose elution. Cramoll 1 was purified by this affinity binding procedure in two steps instead of a previous three-step protocol with ammonium sulfate fractionation, affinity chromatography on Sephadex G-75, and ion exchange chromatography through a CM-cellulose column. PMID:19547713

  2. Affine and deformable registration based on polynomial expansion.

    PubMed

    Farnebäck, Gunnar; Westin, Carl-Fredrik

    2006-01-01

    This paper presents a registration framework based on the polynomial expansion transform. The idea of polynomial expansion is that the image is locally approximated by polynomials at each pixel. Starting with observations of how the coefficients of ideal linear and quadratic polynomials change under translation and affine transformation, algorithms are developed to estimate translation and compute affine and deformable registration between a fixed and a moving image, from the polynomial expansion coefficients. All algorithms can be used for signals of any dimensionality. The algorithms are evaluated on medical data. PMID:17354971

  3. Affinity based and molecularly imprinted cryogels: Applications in biomacromolecule purification.

    PubMed

    Andaç, Müge; Galaev, Igor Yu; Denizli, Adil

    2016-05-15

    The publications in macro-molecularly imprinted polymers have increased drastically in recent years with the development of water-based polymer systems. The macroporous structure of cryogels has allowed the use of these materials within different applications, particularly in affinity purification and molecular imprinting based methods. Due to their high selectivity, specificity, efficient mass transfer and good reproducibility, molecularly imprinted cryogels (MICs) have become attractive for researchers in the separation and purification of proteins. In this review, the recent developments in affinity based cryogels and molecularly imprinted cryogels in protein purification are reviewed comprehensively. PMID:26454622

  4. and as Vertex Operator Extensionsof Dual Affine Algebras

    NASA Astrophysics Data System (ADS)

    Bowcock, P.; Feigin, B. L.; Semikhatov, A. M.; Taormina, A.

    We discover a realisation of the affine Lie superalgebra and of the exceptional affine superalgebra as vertex operator extensions of two algebras with ``dual'' levels (and an auxiliary level-1 algebra). The duality relation between the levels is . We construct the representation of on a sum of tensor products of , , and modules and decompose it into a direct sum over the spectral flow orbit. This decomposition gives rise to character identities, which we also derive. The extension of the construction to is traced to the properties of embeddings into and their relation with the dual pairs. Conversely, we show how the representations are constructed from representations.

  5. Synthesis of biotinylated probes of artemisinin for affinity labeling

    PubMed Central

    Konziase, Benetode

    2015-01-01

    In this data article, we described the synthetic routes to four biotinylated probes (2, 3, 4, and 5) of artemisinin and the associated experimental procedures. We also provided the physical data for the synthesized compounds. These synthesized biotinylated probes of artemisinin are useful molecular tools for the affinity-labeling study of target receptor proteins of artemisinin in tropical pathogens such as Trypanosoma, Leishmania, and Schistosoma. The data provided herein are related to “Biotinylated probes of artemisinin with labeling affinity toward Trypanosoma brucei brucei target proteins”, by Konziase (Anal. Biochem. (2015)). PMID:26217765

  6. Kinetic controlled affinity labeling of target enzyme with thioester chemistry.

    PubMed

    Tomohiro, Takenori; Nakabayashi, Masahiro; Sugita, Yuka; Morimoto, Shota

    2016-08-01

    High specificity has been an important feature in affinity labeling for target profiling. Especially, to label targets via rapidly progressing reactions with consumption of ligand (probe), high specificity of reaction with common functional groups of target protein should be achieved without reactions with similar groups of non-target proteins. Herein, we demonstrate the kinetic controlled affinity labeling of acyl CoA synthetase using a fatty acid analogue containing a phenylthioester linkage. High specificity was attained by accelerating the labeling rate in the binding pocket. This approach could be useful for profiling a series of target enzymes and transporters in signal transduction pathways. PMID:27298000

  7. Affinity Chromatography Purification of Cytochrome c Binding Enzymes

    NASA Astrophysics Data System (ADS)

    Azzi, Angelo; Bill, Kurt; Broger, Clemens

    1982-04-01

    An efficient affinity chromatography procedure for the isolation of mitochondrial cytochrome c oxidase and reductase is described. Saccharomyces cerevisiae cytochrome c was used as a ligand, bound to a thiol-Sepharose 4B gel through cysteine-107. In this way, the site of interaction of cytochrome c with cytochrome oxidase and reductase remained unmodified and available for binding to a number of partner enzymes. The procedure is adequate for the purification of all those proteins having in common the property of binding with high affinity to cytochrome c--e.g., cytochrome c oxidase, reductase, and peroxidase, sulfite oxidase, and reaction centers of photosynthetic bacteria.

  8. Affinity Adsorbents Based on Carriers Activated by Epoxy-compounds

    NASA Astrophysics Data System (ADS)

    Klyashchitskii, B. A.; Kuznetsov, P. V.

    1984-10-01

    The review is devoted to the synthesis and applications of affinity adsorbents based on carriers activated by epoxy-compounds. The methods for the introduction of epoxy-groups into carriers of different chemical types are discussed and conditions for the immobilisation of three-dimensional spacers and low-molecular-weight and polymeric ligands on carriers containing epoxy-groups are considered. Data are presented on the properties and applications of adsorbents of this type in affinity chromatography. The bibliography includes 144 references.

  9. Self-affine analysis of protein energy

    NASA Astrophysics Data System (ADS)

    Figueirêdo, P. H.; Moret, M. A.; Pascutti, P. G.; Nogueira, E.; Coutinho, S.

    2010-07-01

    We study the time series of the total energy of polypeptides and proteins. These time series were generated by molecular dynamics methods and analyzed by applying detrended fluctuation analysis to estimate the long-range power-law correlation, i.e. to measure scaling exponents α. Such exponents were calculated for all systems and their values follow environment conditions, i.e., they are temperature dependent and also, in a continuum medium approach, vary according to the dielectric constants (we simulated ɛ=2 and ɛ=80). The procedure was applied to investigate polyalanines, and other realistic models of proteins (Insect Defensin A and Hemoglobin). The present findings exhibit results that are consistent with previous ones obtained by other methodologies.

  10. Large scale affinity calculations of cyclodextrin host-guest complexes: Understanding the role of reorganization in the molecular recognition process

    PubMed Central

    Wickstrom, Lauren; He, Peng; Gallicchio, Emilio; Levy, Ronald M.

    2013-01-01

    Host-guest inclusion complexes are useful models for understanding the structural and energetic aspects of molecular recognition. Due to their small size relative to much larger protein-ligand complexes, converged results can be obtained rapidly for these systems thus offering the opportunity to more reliably study fundamental aspects of the thermodynamics of binding. In this work, we have performed a large scale binding affinity survey of 57 β-cyclodextrin (CD) host guest systems using the binding energy distribution analysis method (BEDAM) with implicit solvation (OPLS-AA/AGBNP2). Converged estimates of the standard binding free energies are obtained for these systems by employing techniques such as parallel Hamitionian replica exchange molecular dynamics, conformational reservoirs and multistate free energy estimators. Good agreement with experimental measurements is obtained in terms of both numerical accuracy and affinity rankings. Overall, average effective binding energies reproduce affinity rank ordering better than the calculated binding affinities, even though calculated binding free energies, which account for effects such as conformational strain and entropy loss upon binding, provide lower root mean square errors when compared to measurements. Interestingly, we find that binding free energies are superior rank order predictors for a large subset containing the most flexible guests. The results indicate that, while challenging, accurate modeling of reorganization effects can lead to ligand design models of superior predictive power for rank ordering relative to models based only on ligand-receptor interaction energies. PMID:25147485

  11. Students' Understanding of Proportional, Inverse Proportional, and Affine Functions: Two Studies on the Role of External Representations

    ERIC Educational Resources Information Center

    De Bock, Dirk; Van Dooren, Wim; Verschaffel, Lieven

    2015-01-01

    We investigated students' understanding of proportional, inverse proportional, and affine functions and the way this understanding is affected by various external representations. In a first study, we focus on students' ability to model textual descriptions of situations with different kinds of representations of proportional, inverse…

  12. Comparison of Relative Binding Affinities for Trout and Human Estrogen Receptor Based upon Different Competitive Binding Assays

    EPA Science Inventory

    The development of a predictive model based upon a single aquatic species inevitably raises the question of whether this information is valid for other species. To partially address this question, relative binding affinities (RBA) for six alkylphenols (para-substituted, n- and b...

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

    PubMed

    Ball, D J; Nishimura, J S

    1980-11-25

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

  14. Different affinity states of alpha-1 adrenergic receptors defined by agonists and antagonists in bovine aorta plasma membranes

    SciTech Connect

    Jagadeesh, G.; Deth, R.C.

    1987-11-01

    Evidence for a nonlinear relationship between alpha-1 adrenergic receptor occupancy and tissue responses, together with the finding of different affinity states for agonist binding, has raised the possibility of functional heterogeneity of alpha-1 adrenergic receptors. We have conducted studies to examine: 1) binding characteristics of (/sup 3/H)prazosin, 2) competition of antagonists at these sites and 3) different affinity states of the receptor for agonists and modulation of these states by 5'-guanylylimidodiphosphate (Gpp(NH)p). A plasma membrane-enriched vesicular fraction (F2; 15%/33% sucrose interphase) was prepared from the muscular medial layer of bovine thoracic aorta. (/sup 3/H)Prazosin binding was characterized by a monophasic saturation isotherm (KD = 0.116 nM, Bmax = 112 fmol/mg of protein). Antagonist displacement studies yielded a relative potency order of prazosin greater than or equal to WB4104 much greater than phentolamine greater than corynanthine greater than yohimbine greater than or equal to idazoxan greater than rauwolscine. Competition curves for unlabeled prazosin, WB4101 (2-(2,6-dimethoxyphenoxyethyl)-aminomethyl-1,4 benzodioxane) and phentolamine were shallow and were best modeled to two binding sites with picomolar and nanomolar KD values. Gpp(NH)p was without effect on antagonist affinity. Agonist (epinephrine, norepinephrine and phenylephrine) competition with (/sup 3/H)prazosin binding was biphasic with pseudo-Hill slopes less than 1.0. Binding was best described by a two-site model in which the average contribution of high affinity sites was 23% of total binding. KD values for the high affinity site ranged from 2.9 to 18 nM, and 3.9 to 5.0 microM for the low affinity site.

  15. Biotinylated Cyclophane: Synthesis, Cyclophane-Avidin Conjugates, and Their Enhanced Guest-Binding Affinity.

    PubMed

    Hayashida, Osamu; Kojima, Miwa; Kusano, Shuhei

    2015-10-01

    Cationic and anionic cyclophanes bearing a biotin moiety were synthesized as a water-soluble host (1a and 1b, respectively). Both hosts 1a and 1b were found to strongly bind avidin with binding constants of 1.3 × 10(8) M(-1), as confirmed by surface plasmon resonance measurements. The present conjugate of 1a with avidin (1a-avidin) showed an enhanced guest binding affinity toward fluorescence guests such as TNS and 2,6-ANS. The K values of 1a-avidin conjugate with TNS and 2,6-ANS were ~19-fold larger than those of monocyclic cyclophane 1a with the identical guests. Favorable hydrophobic and electrostatic interactions between 1a-avidin and TNS were suggested by computer-aided molecular modeling calculations. Moreover, addition of excess biotin to the complexes of 1a-avidin with the guests resulted in dissociation of 1a-avidin to avidin and 1a having less guest-binding affinity. Conversely, such enhancements in the guest-binding affinity were not obviously observed for the conjugate of anionic 1b with avidin (1b-avidin) due to electrostatic repulsion between anionic 1b and anionic guests. PMID:26360807

  16. Modulation of Perfusion and Oxygenation by Red Blood Cell Oxygen Affinity during Acute Anemia

    PubMed Central

    Cabrales, Pedro; Tsai, Amy G.; Intaglietta, Marcos

    2008-01-01

    Responses to exchange transfusion using red blood cells (RBCs) with modified hemoglobin (Hb) oxygen (O2) affinity were studied in the hamster window chamber model during acute anemia to determine its role on microvascular perfusion and tissue oxygenation. Allosteric effectors were introduced in the RBCs by electroporation. Inositol hexaphosphate (IHP) and 5-hydroxymethyl-2-furfural (5HMF) were used to decrease and increase Hb-O2 affinity. In vitro P50s (partial pressure of O2 at 50% Hb saturation) were modified to 10, 25, 45, and 50 mm Hg (normal P50 is 32 mm Hg). Allosteric effectors also decreased the Hill coefficient. Anemic condition was induced by isovolemic hemodilution exchanges using 6% dextran 70 kD to 18% hematocrit (Hct). Modified RBCs (at 18% Hct in 5% albumin solution) were infused by exchange transfusion of 35% of blood volume. Systemic parameters, microvascular perfusion, capillary perfusion (functional capillary density, FCD), and microvascular Po2 levels were measured. RBcs with P50 of 45 mm Hg increased tissue Po2 and decreased O2 delivery (Do2) and extraction (Vo2) and RBCs with P50 of 60 mmHg reduced FCD, microvascular flow, tissue Po2, Do2 and Vo2. Erythrocytes with increased Hb-O2 affinity maintained hemodynamic conditions, Do2 and decreased tissue Po2. This study shows that in an anemic condition, maximal tissue Po2 does not correspond to maximal Do2 and Vo2. PMID:17884988

  17. Deconvolution of antibody affinities and concentrations by non-linear regression analysis of competitive ELISA data.

    SciTech Connect

    Stevens, F. J.; Bobrovnik, S. A.; Biosciences Division; Palladin Inst. Biochemistry

    2007-12-01

    Physiological responses of the adaptive immune system are polyclonal in nature whether induced by a naturally occurring infection, by vaccination to prevent infection or, in the case of animals, by challenge with antigen to generate reagents of research or commercial significance. The composition of the polyclonal responses is distinct to each individual or animal and changes over time. Differences exist in the affinities of the constituents and their relative proportion of the responsive population. In addition, some of the antibodies bind to different sites on the antigen, whereas other pairs of antibodies are sterically restricted from concurrent interaction with the antigen. Even if generation of a monoclonal antibody is the ultimate goal of a project, the quality of the resulting reagent is ultimately related to the characteristics of the initial immune response. It is probably impossible to quantitatively parse the composition of a polyclonal response to antigen. However, molecular regression allows further parameterization of a polyclonal antiserum in the context of certain simplifying assumptions. The antiserum is described as consisting of two competing populations of high- and low-affinity and unknown relative proportions. This simple model allows the quantitative determination of representative affinities and proportions. These parameters may be of use in evaluating responses to vaccines, to evaluating continuity of antibody production whether in vaccine recipients or animals used for the production of antisera, or in optimizing selection of donors for the production of monoclonal antibodies.

  18. Assessment of Binding Affinity between Drugs and Human Serum Albumin Using Nanoporous Anodic Alumina Photonic Crystals.

    PubMed

    Nemati, Mahdieh; Santos, Abel; Law, Cheryl Suwen; Losic, Dusan

    2016-06-01

    In this study, we report an innovative approach aiming to assess the binding affinity between drug molecules and human serum albumin by combining nanoporous anodic alumina rugate filters (NAA-RFs) modified with human serum albumin (HSA) and reflectometric interference spectroscopy (RIfS). NAA-RFs are photonic crystal structures produced by sinusoidal pulse anodization of aluminum that present two characteristic optical parameters, the characteristic reflection peak (λPeak), and the effective optical thickness of the film (OTeff), which can be readily used as sensing parameters. A design of experiments strategy and an ANOVA analysis are used to establish the effect of the anodization parameters (i.e., anodization period and anodization offset) on the sensitivity of HSA-modified NAA-RFs toward indomethacin, a model drug. To this end, two sensing parameters are used, that is, shifts in the characteristic reflection peak (ΔλPeak) and changes in the effective optical thickness of the film (ΔOTeff). Subsequently, optimized NAA-RFs are used as sensing platforms to determine the binding affinity between a set of drugs (i.e., indomethacin, coumarin, sulfadymethoxine, warfarin, and salicylic acid) and HSA molecules. Our results verify that the combination of HSA-modified NAA-RFs with RIfS can be used as a portable, low-cost, and simple system for establishing the binding affinity between drugs and plasma proteins, which is a critical factor to develop efficient medicines for treating a broad range of diseases and medical conditions. PMID:27128744

  19. Oligomerization of Peptides LVEALYL and RGFFYT and Their Binding Affinity to Insulin

    PubMed Central

    Chiang, Hsin-Lin; Ngo, Son Tung; Chen, Chun-Jung; Hu, Chin-Kun; Li, Mai Suan

    2013-01-01

    Recently it has been proposed a model for fibrils of human insulin in which the fibril growth proceeds via stacking LVEALYL (fragment 11–17 from chain B of insulin) into pairs of tightly interdigitated -sheets. The experiments have also shown that LVEALYL has high propensity to self-assembly and binding to insulin. This necessitates study of oligomerization of LVEALYL and its binding affinity to full-length insulin. Using the all-atom simulations with Gromos96 43a1 force field and explicit water it is shown that LVEALYL can aggregate. Theoretical estimation of the binding free energy of LVEALYL to insulin by the molecular mechanic Poisson-Boltzmann surface area method reveals its strong binding affinity to chain B, implying that, in agreement with the experiments, LVEALYL can affect insulin aggregation via binding mechanism. We predict that, similar to LVEALYL, peptide RGFFYT (fragment B22-27) can self-assemble and bind to insulin modulating its fibril growth process. The binding affinity of RGFFYT is shown to be comparable with that of LVEALYL. PMID:23805182

  20. Predicting the Impact of Missense Mutations on Protein-Protein Binding Affinity.

    PubMed

    Li, Minghui; Petukh, Marharyta; Alexov, Emil; Panchenko, Anna R

    2014-04-01

    The crucial prerequisite for proper biological function is the protein's ability to establish highly selective interactions with macromolecular partners. A missense mutation that alters the protein binding affinity may cause significant perturbations or complete abolishment of the function, potentially leading to diseases. The availability of computational methods to evaluate the impact of mutations on protein-protein binding is critical for a wide range of biomedical applications. Here, we report an efficient computational approach for predicting the effect of single and multiple missense mutations on protein-protein binding affinity. It is based on a well-tested simulation protocol for structure minimization, modified MM-PBSA and statistical scoring energy functions with parameters optimized on experimental sets of several thousands of mutations. Our simulation protocol yields very good agreement between predicted and experimental values with Pearson correlation coefficients of 0.69 and 0.63 and root-mean-square errors of 1.20 and 1.90 kcal mol(-1) for single and multiple mutations, respectively. Compared with other available methods, our approach achieves high speed and prediction accuracy and can be applied to large datasets generated by modern genomics initiatives. In addition, we report a crucial role of water model and the polar solvation energy in estimating the changes in binding affinity. Our analysis also reveals that prediction accuracy and effect of mutations on binding strongly depends on the type of mutation and its location in a protein complex. PMID:24803870

  1. Fluorescent Boronic Acid Polymer Grafted on Silica Particles for Affinity Separation of Saccharides

    PubMed Central

    2014-01-01

    Boronic acid affinity gels are important for effective separation of biological active cis-diols, and are finding applications both in biotech industry and in biomedical research areas. To increase the efficacy of boronate affinity separation, it is interesting to introduce repeating boronic acid units in flexible polymer chains attached on solid materials. In this work, we synthesize polymer brushes containing boronic acid repeating units on silica gels using surface-initiated atom transfer radical polymerization (ATRP). A fluorescent boronic acid monomer is first prepared from an azide-tagged fluorogenic boronic acid and an alkyne-containing acrylate by Cu(I)-catalyzed 1,3-dipolar cycloaddition reaction (the CuAAC click chemistry). The boronic acid monomer is then grafted to the surface of silica gel modified with an ATRP initiator. The obtained composite material contains boronic acid polymer brushes on surface and shows favorable saccharide binding capability under physiological pH conditions, and displays interesting fluorescence intensity change upon binding fructose and glucose. In addition to saccharide binding, the flexible polymer brushes on silica also enable fast separation of a model glycoprotein based on selective boronate affinity interaction. The synthetic approach and the composite functional material developed in this work should open new opportunities for high efficiency detection, separation, and analysis of not only simple saccharides, but also glycopeptides and large glycoproteins. PMID:24444898

  2. Choline uptake in Agrobacterium tumefaciens by the high-affinity ChoXWV transporter.

    PubMed

    Aktas, Meriyem; Jost, Kathinka A; Fritz, Christiane; Narberhaus, Franz

    2011-10-01

    Agrobacterium tumefaciens is a facultative phytopathogen that causes crown gall disease. For successful plant transformation A. tumefaciens requires the membrane lipid phosphatidylcholine (PC), which is produced via the methylation and the PC synthase (Pcs) pathways. The latter route is dependent on choline. Although choline uptake has been demonstrated in A. tumefaciens, the responsible transporter(s) remained elusive. In this study, we identified the first choline transport system in A. tumefaciens. The ABC-type choline transporter is encoded by the chromosomally located choXWV operon (ChoX, binding protein; ChoW, permease; and ChoV, ATPase). The Cho system is not critical for growth and PC synthesis. However, [14C]choline uptake is severely reduced in A. tumefaciens choX mutants. Recombinant ChoX is able to bind choline with high affinity (equilibrium dissociation constant [KD] of ≈2 μM). Since other quaternary amines are bound by ChoX with much lower affinities (acetylcholine, KD of ≈80 μM; betaine, KD of ≈470 μM), the ChoXWV system functions as a high-affinity transporter with a preference for choline. Two tryptophan residues (W40 and W87) located in the predicted ligand-binding pocket are essential for choline binding. The structural model of ChoX built on Sinorhizobium meliloti ChoX resembles the typical structure of substrate binding proteins with a so-called "Venus flytrap mechanism" of substrate binding. PMID:21803998

  3. Method for trapping affinity chromatography of transcription factors using aldehyde-hydrazide coupling to agarose.

    PubMed

    Jia, Yinshan; Jarrett, Harry W

    2015-08-01

    The use of a method of coupling DNA was investigated for trapping and purifying transcription factors. Using the GFP-C/EBP (CAAT/enhancer binding protein) fusion protein as a model, trapping gives higher purity and comparable yield to conventional affinity chromatography. The chemistry used is mild and was shown to have no detrimental effect on GFP fluorescence or GFP-C/EBP DNA binding. The method involves introducing a ribose nucleotide to the 3' end of a DNA sequence. Reaction with mM NaIO4 (sodium metaperiodate) produces a dialdehyde of ribose that couples to hydrazide-agarose. The DNA is combined at nM concentration with a nuclear extract or other protein mixture, and DNA-protein complexes form. The complex is then coupled to hydrazide-agarose for trapping the DNA-protein complex and the protein eluted by increasing NaCl concentration. Using a different oligonucleotide with the proximal E-box sequence from the human telomerase promoter, USF-2 transcription factor was purified by trapping, again with higher purity than results from conventional affinity chromatography and similar yield. Other transcription factors binding E-boxes, including E2A, c-Myc, and Myo-D, were also purified, but myogenin and NFκB were not. Therefore, this approach proved to be valuable for both affinity chromatography and the trapping approach. PMID:25935261

  4. Method for trapping affinity chromatography of transcription factors using aldehyde-hydrazide coupling to agarose

    PubMed Central

    Jia, Yinshan; Jarrett, Harry W.

    2015-01-01

    The uses of a method of coupling DNA is investigated for trapping and purifying transcription factors. Using the GFP-C/EBP fusion protein as a model, trapping gives higher purity and comparable yield to conventional affinity chromatography. The chemistry utilized is mild and was shown to have no detrimental effect on GFP fluorescence or GFP-C/EBP DNA-binding. The method involves introducing a ribose nucleotide to the 3′ end of a DNA sequence. Reaction with mM NaIO4 (sodium metaperiodate) produces a dialdehyde of ribose which couples to hydrazide-agarose. The DNA is combined at nM concentration with a nuclear extract or other protein mixture and DNA-protein complexes form. The complex is then coupled to hydrazide-agarose for trapping the DNA-protein complex and the protein eluted by increasing NaCl concentration. Using a different oligonucleotide with the proximal E-box sequence from the human telomerase promoter, USF-2 transcription factor was purified by trapping, again with higher purity than results from conventional affinity chromatography and similar yield. Other transcription factors binding E-boxes including E2A, c-myc, and myo-D were also purified but myogenenin and NFκB were not. Therfore, this approach proved valuable for both affinity chromatography and for the trapping approach. PMID:25935261

  5. Fluorescent boronic acid polymer grafted on silica particles for affinity separation of saccharides.

    PubMed

    Xu, Zhifeng; Uddin, Khan Mohammad Ahsan; Kamra, Tripta; Schnadt, Joachim; Ye, Lei

    2014-02-12

    Boronic acid affinity gels are important for effective separation of biological active cis-diols, and are finding applications both in biotech industry and in biomedical research areas. To increase the efficacy of boronate affinity separation, it is interesting to introduce repeating boronic acid units in flexible polymer chains attached on solid materials. In this work, we synthesize polymer brushes containing boronic acid repeating units on silica gels using surface-initiated atom transfer radical polymerization (ATRP). A fluorescent boronic acid monomer is first prepared from an azide-tagged fluorogenic boronic acid and an alkyne-containing acrylate by Cu(I)-catalyzed 1,3-dipolar cycloaddition reaction (the CuAAC click chemistry). The boronic acid monomer is then grafted to the surface of silica gel modified with an ATRP initiator. The obtained composite material contains boronic acid polymer brushes on surface and shows favorable saccharide binding capability under physiological pH conditions, and displays interesting fluorescence intensity change upon binding fructose and glucose. In addition to saccharide binding, the flexible polymer brushes on silica also enable fast separation of a model glycoprotein based on selective boronate affinity interaction. The synthetic approach and the composite functional material developed in this work should open new opportunities for high efficiency detection, separation, and analysis of not only simple saccharides, but also glycopeptides and large glycoproteins. PMID:24444898

  6. Optimal fusion of antibody binding domains resulted in higher affinity and wider specificity.

    PubMed

    Dong, Jinhua; Kojima, Tomoki; Ohashi, Hiroyuki; Ueda, Hiroshi

    2015-11-01

    Antibody is a very important protein in biotechnological and biomedical fields because of its high affinity and specificity to various antigens. Due to the rise of human antibody therapeutics, its cost-effective purification is an urgent issue for bio-industry. In this study, we made novel fusion proteins PAxPG with a flexible (DDAKK)n linker between the two Ig binding domains derived from Staphylococcus protein A and Streptococcus protein G. The fusion proteins bound human and mouse IgGs and their fragments with up to 58-times higher affinity and wider specificity than the parental binding domains. Interestingly, the optimal linker for human Fab fragment was n = 4, which was close to the modeled distance between the termini of domains bound to heavy chain, implying increased avidity as a possible mechanism. For binding to Fc, the longest n=6 linker gave the highest affinity, implying longer interchain distance between the two binding sites. The novel fusion protein with optimized interdomain linker length will be a useful tool for the purification and detection of various IgGs including mouse IgG1 that binds only weakly to natural protein A. PMID:25910963

  7. Weak affinity chromatography as a new approach for fragment screening in drug discovery.

    PubMed

    Duong-Thi, Minh-Dao; Meiby, Elinor; Bergström, Maria; Fex, Tomas; Isaksson, Roland; Ohlson, Sten

    2011-07-01

    Fragment-based drug design (FBDD) is currently being implemented in drug discovery, creating a demand for developing efficient techniques for fragment screening. Due to the intrinsic weak or transient binding of fragments (mM-μM in dissociation constant (K(D))) to targets, methods must be sensitive enough to accurately detect and quantify an interaction. This study presents weak affinity chromatography (WAC) as an alternative tool for screening of small fragments. The technology was demonstrated by screening of a selected 23-compound fragment collection of documented binders, mostly amidines, using trypsin and thrombin as model target protease proteins. WAC was proven to be a sensitive, robust, and reproducible technique that also provides information about affinity of a fragment in the range of 1 mM-10 μM. Furthermore, it has potential for high throughput as was evidenced by analyzing mixtures in the range of 10 substances by WAC-MS. The accessibility and flexibility of the technology were shown as fragment screening can be performed on standard HPLC equipment. The technology can further be miniaturized and adapted to the requirements of affinity ranges of the fragment library. All these features of WAC make it a potential method in drug discovery for fragment screening. PMID:21352794

  8. Molecular-size-dependent variations in the proportions of chains with high binding affinities for antithrombin in rat skin heparin proteoglycans.

    PubMed Central

    Horner, A A

    1989-01-01

    Approximately half of all rat skin heparin proteoglycans have polysaccharide chains that have no sites with high binding affinity for antithrombin. The rest have chains with high-affinity antithrombin-binding-site densities ranging from zero to five sites per chain, with a high degree of variation. Proteoglycans vary in size because of diversity in the number of chains per molecule; the relationship between proteoglycan size and high-affinity antithrombin-binding-site density has not been studied previously. Polydisperse heparin proteoglycans from rat skin, labelled biosynthetically with 35S, were fractionated by gel filtration on Bio-Gel A-150m and arbitrarily divided into five fractions of decreasing average molecular size. Fractionation of these products on antithrombin-agarose showed that the proportion of proteoglycans with high affinity for antithrombin decreased from 39% to 25% as molecular size decreased. However, as the molecular size of high-affinity proteoglycans decreased, the proportion of their chains that had high affinity increased from 29% to 59%. Therefore molecular size is a significant factor in determining the proportion of high-affinity chains in heparin proteoglycans. A model of heparin biosynthesis is proposed in which areas of specific enzyme activity that control the synthesis of the antithrombin-binding-site sequence are sparsely and nonrandomly distributed on mast-cell Golgi membranes. It is postulated that the likelihood of a developing proteoglycan encountering one of these hypothetical areas is molecular-size-dependent. Images Fig. 1. PMID:2590178

  9. Phosphatidylserine and Phosphatidylethanolamine Bind to Protein Z Cooperatively and with Equal Affinity.

    PubMed

    Sengupta, Tanusree; Manoj, Narayanan

    2016-01-01

    Protein Z (PZ) is an anticoagulant that binds with high affinity to Protein Z-dependent protease inhibitor (ZPI) and accelerates the rate of ZPI-mediated inhibition of factor Xa (fXa) by more than 1000-fold in the presence of Ca2+ and phospholipids. PZ promotion of the ZPI-fXa interaction results from the anchoring of the Gla domain of PZ onto phospholipid surfaces and positioning the bound ZPI in close proximity to the Gla-anchored fXa, forming a ternary complex of PZ/ZPI/fXa. Although interaction of PZ with phospholipid membrane appears to be absolutely crucial for its cofactor activity, little is known about the binding of different phospholipids to PZ. The present study was conceived to understand the interaction of different phospholipids with PZ. Experiments with both soluble lipids and model membranes revealed that PZ binds to phosphatidylserine (PS) and phosphatidylethanolamine (PE) with equal affinity (Kd~48 μM); further, PS and PE bound to PZ synergistically. Equilibrium dialysis experiments revealed two lipid-binding sites for both PS and PE. PZ binds with weaker affinity to other phospholipids, e.g., phosphatidic acid, phosphatidylglycerol, phosphatidylcholine and binding of these lipids is not synergistic with respect to PS. Both PS and PE -containing membranes supported the formation of a fXa-PZ complex. PZ protection of fXa from antithrombin inhibition were also shown to be comparable in presence of both PS: PC and PE: PC membranes. These findings are particularly important and intriguing since they suggest a special affinity of PZ, in vivo, towards activated platelets, the primary membrane involved in blood coagulation process. PMID:27584039

  10. Insights into the conformational equilibria of maltose-binding protein by analysis of high affinity mutants.

    PubMed

    Telmer, Patrick G; Shilton, Brian H

    2003-09-01

    The affinity of maltose-binding protein (MBP) for maltose and related carbohydrates was greatly increased by removal of groups in the interface opposite the ligand binding cleft. The wild-type protein has a KD of 1200 nM for maltose; mutation of residues Met-321 and Gln-325, both to alanine, resulted in a KD for maltose of 70 nM; deletion of 4 residues, Glu-172, Asn-173, Lys-175, and Tyr-176, which are part of a poorly ordered loop, results in a KD for maltose of 110 nM. Combining the mutations yields an increased affinity for maltodextrins and a KD of 6 nM for maltotriose. Comparison of ligand binding by the mutants, using surface plasmon resonance spectroscopy, indicates that decreases in the off-rate are responsible for the increased affinity. Small-angle x-ray scattering was used to demonstrate that the mutations do not significantly affect the solution conformation of MBP in either the presence or absence of maltose. The crystal structures of selected mutants showed that the mutations do not cause significant structural changes in either the closed or open conformation of MBP. These studies show that interactions in the interface opposite the ligand binding cleft, which we term the "balancing interface," are responsible for modulating the affinity of MBP for its ligand. Our results are consistent with a model in which the ligand-bound protein alternates between the closed and open conformations, and removal of interactions in the balancing interface decreases the stability of the open conformation, without affecting the closed conformation. PMID:12794084

  11. Affinity purification of recombinant proteins using a novel silica-binding peptide as a fusion tag.

    PubMed

    Abdelhamid, Mohamed A A; Motomura, Kei; Ikeda, Takeshi; Ishida, Takenori; Hirota, Ryuichi; Kuroda, Akio

    2014-06-01

    We recently reported that silica is deposited on the coat of Bacillus cereus spores as a layer of nanometer-sized particles (Hirota et al. 2010 J Bacteriol 192: 111-116). Gene disruption analysis revealed that the spore coat protein CotB1 mediates the accumulation of silica (our unpublished results). Here, we report that B. cereus CotB1 (171 amino acids [aa]) and its C-terminal 14-aa region (corresponding to residues 158-171, designated CotB1p) show strong affinity for silica particles, with dissociation constants at pH 8.0 of 2.09 and 1.24 nM, respectively. Using CotB1 and CotB1p as silica-binding tags, we developed a silica-based affinity purification method in which silica particles are used as an adsorbent for CotB1/CotB1p fusion proteins. Small ubiquitin-like modifier (SUMO) technology was employed to release the target proteins from the adsorbed fusion proteins. SUMO-protease-mediated site-specific cleavage at the C-terminus of the fused SUMO sequence released the tagless target proteins into the liquid phase while leaving the tag region still bound to the solid phase. Using the fluorescent protein mCherry as a model, our purification method achieved 85 % recovery, with a purity of 95 % and yields of 0.60 ± 0.06 and 1.13 ± 0.13 mg per 10-mL bacterial culture for the CotB1-SUMO-mCherry and CotB1p-SUMO-mCherry fusions, respectively. CotB1p, a short 14-aa peptide, which demonstrates high affinity for silica, could be a promising fusion tag for both affinity purification and enzyme immobilization on silica supports. PMID:24756322

  12. Engineering high-affinity PD-1 variants for optimized immunotherapy and immuno-PET imaging

    PubMed Central

    Maute, Roy L.; Gordon, Sydney R.; Mayer, Aaron T.; McCracken, Melissa N.; Natarajan, Arutselvan; Ring, Nan Guo; Kimura, Richard; Tsai, Jonathan M.; Manglik, Aashish; Kruse, Andrew C.; Gambhir, Sanjiv S.; Weissman, Irving L.; Ring, Aaron M.

    2015-01-01

    Signaling through the immune checkpoint programmed cell death protein-1 (PD-1) enables tumor progression by dampening antitumor immune responses. Therapeutic blockade of the signaling axis between PD-1 and its ligand programmed cell death ligand-1 (PD-L1) with monoclonal antibodies has shown remarkable clinical success in the treatment of cancer. However, antibodies have inherent limitations that can curtail their efficacy in this setting, including poor tissue/tumor penetrance and detrimental Fc-effector functions that deplete immune cells. To determine if PD-1:PD-L1–directed immunotherapy could be improved with smaller, nonantibody therapeutics, we used directed evolution by yeast-surface display to engineer the PD-1 ectodomain as a high-affinity (110 pM) competitive antagonist of PD-L1. In contrast to anti–PD-L1 monoclonal antibodies, high-affinity PD-1 demonstrated superior tumor penetration without inducing depletion of peripheral effector T cells. Consistent with these advantages, in syngeneic CT26 tumor models, high-affinity PD-1 was effective in treating both small (50 mm3) and large tumors (150 mm3), whereas the activity of anti–PD-L1 antibodies was completely abrogated against large tumors. Furthermore, we found that high-affinity PD-1 could be radiolabeled and applied as a PET imaging tracer to efficiently distinguish between PD-L1–positive and PD-L1–negative tumors in living mice, providing an alternative to invasive biopsy and histological analysis. These results thus highlight the favorable pharmacology of small, nonantibody therapeutics for enhanced cancer immunotherapy and immune diagnostics. PMID:26604307

  13. Qualitative cues in the discrimination of affine-transformed minimal patterns.

    PubMed

    Kukkonen, H T; Foster, D H; Wood, J R; Wagemans, J; Van Gool, L

    1996-01-01

    An important factor in judging whether two retinal images arise from the same object viewed from different positions may be the presence of certain properties or cues that are 'qualitative invariants' with respect to the natural transformations, particularly affine transformations, associated with changes in viewpoint. To test whether observers use certain affine qualitative cues such as concavity, convexity, collinearity, and parallelism of the image elements, a 'same-different' discrimination experiment was carried out with planar patterns that were defined by four points either connected by straight line segments (line patterns) or marked by dots (dot patterns). The first three points of each pattern were generated randomly; the fourth point fell on their diagonal bisector. According to the position of that point, the patterns were concave, triangular (three points being collinear), convex, or parallel sided. In a 'same' trial, an affine transformation was applied to one of two identical patterns; in a 'different' trial, the affine transformation was applied after the point lying on the diagonal bisector was perturbed a short, fixed distance along the bisector, inwards for one pattern and outwards for the other. Observers' ability to discriminate 'same' from 'different' pairs of patterns depended strongly on the position of the fourth, displaced, point: performance varied rapidly when the position of the displaced point was such that the patterns were nearly triangular or nearly parallel sided, consistent with observers using the hypothesised qualitative cues. The experimental data were fitted with a simple probabilistic model of discrimination performance that used a combination of these qualitative cues and a single quantitative cue. PMID:8733148

  14. Enhanced antigen-antibody binding affinity mediated by an anti-idiotypic antibody

    SciTech Connect

    Sawutz, D.G.; Koury, R.; Homcy, C.J.

    1987-08-25

    The authors previously described the production of four monoclonal antibodies to the ..beta..-adrenergic receptor antagonist alprenolol. One of these antibodies, 5B7 (IgG/sub 2a/, kappa), was used to raise anti-idiotypic antisera in rabbits. In contrast to the expected results, one of the anti-idiotypic antisera (R9) promotes (/sup 125/I)iodocyanopinodolol (ICYP) binding to antibody 5B7. In the presence of R9, the dissociation constant decreases 100-fold from 20 to 0.3 nM. This increase in binding affinity of antibody 5B7 for ICYP is not observed in the presence of preimmune, rabbit anti-mouse or anti-idiotypic antisera generated to a monoclonal antibody of a different specificity. Furthermore, R9 in the absence of 5B7 does not bind ICYP. The F(ab) fragments of 5B7 and T9 behaved in a similar manner, and the soluble complex responsible for the high-affinity interaction with ICYP can be identified by gel filtration chromatography. The elution position of the complex is consistent with a 5B7 F(ab)-R9 F(ab) dimer, indicating that polyvalency is not responsible for the enhanced ligand binding. Kinetic analysis of ICYP-5B7 binding revealed that the rate of ICYP dissociation from 5B7 in the presence of R9 is approximately 100 times slower than in the absence of R9, consistent with the 100-fold change in binding affinity of 5B7 for ICYP. The available data best fit a model in which an anti-idiotypic antibody binds at or near the binding site of the idiotype participating in the formation of a hybrid ligand binding site. This would allow increased contact of the ligand with the idiotype-anti-idiotype complex and result in an enhanced affinity of the ligand interaction.

  15. Quantitative demonstration of intrathecal synthesis of high affinity immunoglobulin G in herpes simplex encephalitis using affinity-mediated immunoblotting.

    PubMed

    Chapman, Miles D; Thompson, Edward J; Candler, Paul M; Dale, Russell C; Church, Andrew J; Giovannoni, Gavin

    2007-04-01

    Three paired serial samples of CSF and serum (from days 8, 13 and 22) were taken from a patient referred to the National Hospital for Neurology and Neurosurgery with what was duly confirmed as having herpes simplex encephalitis using PCR. The samples were investigated using affinity-mediated immunoblotting followed by incubation with sodium thiocyanate. Digitisation of the blots enabled further analysis. We showed that the clones of antigen-specific IgG, which were produced intrathecally, were of higher relative affinity than polyclonal antigen-specific IgG. PMID:17303253

  16. Electrochemical affinity biosensors for detection of mycotoxins: A review.

    PubMed

    Vidal, Juan C; Bonel, Laura; Ezquerra, Alba; Hernández, Susana; Bertolín, Juan R; Cubel, Carlota; Castillo, Juan R

    2013-11-15

    This review discusses the current state of electrochemical biosensors in the determination of mycotoxins in foods. Mycotoxins are highly toxic secondary metabolites produced by molds. The acute toxicity of these results in serious human and animal health problems, although it has been only since early 1960s when the first studied aflatoxins were found to be carcinogenic. Mycotoxins affect a broad range of agricultural products, most important cereals and cereal-based foods. A majority of countries, mentioning especially the European Union, have established preventive programs to control contamination and strict laws of the permitted levels in foods. Official methods of analysis of mycotoxins normally requires sophisticated instrumentation, e.g. liquid chromatography with fluorescence or mass detectors, combined with extraction procedures for sample preparation. For about sixteen years, the use of simpler and faster analytical procedures based on affinity biosensors has emerged in scientific literature as a very promising alternative, particularly electrochemical (i.e., amperometric, impedance, potentiometric or conductimetric) affinity biosensors due to their simplicity and sensitivity. Typically, electrochemical biosensors for mycotoxins use specific antibodies or aptamers as affinity ligands, although recombinant antibodies, artificial receptors and molecular imprinted polymers show potential utility. This article deals with recent advances in electrochemical affinity biosensors for mycotoxins and covers complete literature from the first reports about sixteen years ago. PMID:23743326

  17. Affinities and beyond! Developing Ways of Seeing in Online Spaces

    ERIC Educational Resources Information Center

    Davies, Julia

    2006-01-01

    This article presents an insider view of an online community of adults involved in sharing digital photography through a host website, Flickr. It describes how reciprocal teaching and learning partnerships in a dynamic multimodal environment are achieved through the creation of a "Third Space" or "Affinity Space", where "Funds of Knowledge" are…

  18. Native Elution of Yeast Protein Complexes Obtained by Affinity Capture.

    PubMed

    LaCava, John; Fernandez-Martinez, Javier; Rout, Michael P

    2016-01-01

    This protocol describes two options for the native (nondenaturing) elution of protein complexes obtained by affinity capture. The first approach involves the elution of complexes purified through a tag that includes a human rhinovirus 3C protease (PreScission protease) cleavage site sequence between the protein of interest and the tag. Incubation with the protease cleaves immobilized complexes from the affinity medium. The second approach involves the release of protein A-tagged protein complexes using a competitive elution reagent called PEGylOx. The degree of purity of the native assemblies eluted is sample dependent and strongly influenced by the affinity capture. It should be noted that the efficiency of native elution is commonly lower than that of elution by a denaturing agent (e.g., SDS) and the release of the complex will be limited by the activity of the protease or the inhibition constant (Ki) of the competitive release agent. However, an advantage of native release is that some nonspecifically bound materials tend to stay adsorbed to the affinity medium, providing an eluted fraction of higher purity. Finally, keep in mind that the presence of the protease or elution peptide could potentially affect downstream applications; thus, their removal should be considered. PMID:27371597

  19. Affinity of cefoperazone for penicillin-binding proteins.

    PubMed Central

    Matsubara, N; Minami, S; Matsuhashi, M; Takaoka, M; Mitsuhashi, S

    1980-01-01

    Cefoperazone (T-1551, CFP) a new semisynthetic cephalosporin, has a broad spectrum of antibacterial activity. We investigated the affinity of CFP to penicillin-binding proteins (PBPs) and the inhibition of peptidoglycan synthesis by CFP. CFP had high affinities for Escherichia coli PBP-3, -1Bs, -2, and -1A, in descending order, and low affinities for PBP-4, -5, and -6. Similarly, CFP showed high affinity for Pseudomonas aeruginosa PBP-3, -1A, -1B, -2, and -4, in descending order. It is known that E. coli PBP-3 and P. aeruginosa PBP-3 participate in cell division. These results are in good agreement with the formation of filamentous cells of E. coli and P. aeruginosa treated with CFP. CFP had lower inhibitory activities on D-alanine carboxypeptidase IA and IB of E. coli than that of penicillin G, but its inhibitory activities on the cross-link formation in peptidoglycan synthesis were the same as those of penicillin G and higher than those of ampicillin. Images PMID:6448021

  20. Development of gadolinium based nanoparticles having an affinity towards melanin

    NASA Astrophysics Data System (ADS)

    Morlieras, Jessica; Chezal, Jean-Michel; Miot-Noirault, Elisabeth; Roux, Amandine; Heinrich-Balard, Laurence; Cohen, Richard; Tarrit, Sébastien; Truillet, Charles; Mignot, Anna; Hachani, Roxanne; Kryza, David; Antoine, Rodolphe; Dugourd, Philippe; Perriat, Pascal; Janier, Marc; Sancey, Lucie; Lux, François; Tillement, Olivier

    2013-01-01

    Small Rigid Platforms (SRPs) are sub-5 nanometre gadolinium based nanoparticles that have been developed for multimodal imaging and theranostic applications. They are composed of a polysiloxane network surrounded by gadolinium chelates. A covalent coupling with quinoxaline derivatives has been performed. Such derivatives have proven their affinity for melanin frequently expressed in primary melanoma cases. Three different quinoxaline derivatives have been synthesised and coupled to the nanoparticles. The affinity of the grafted nanoparticles for melanin has then been shown in vitro by surface plasmon resonance on a homemade melanin grafted gold chip.Small Rigid Platforms (SRPs) are sub-5 nanometre gadolinium based nanoparticles that have been developed for multimodal imaging and theranostic applications. They are composed of a polysiloxane network surrounded by gadolinium chelates. A covalent coupling with quinoxaline derivatives has been performed. Such derivatives have proven their affinity for melanin frequently expressed in primary melanoma cases. Three different quinoxaline derivatives have been synthesised and coupled to the nanoparticles. The affinity of the grafted nanoparticles for melanin has then been shown in vitro by surface plasmon resonance on a homemade melanin grafted gold chip. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr33457g

  1. Affinity through Mathematical Activity: Cultivating Democratic Learning Communities

    ERIC Educational Resources Information Center

    Sengupta-Irving, Tesha

    2014-01-01

    In this article, the author demonstrates how a broader view of what shapes affinity is ideologically and practically linked to creating democratic learning communities. Specifically, the author explores how a teacher employed complex instruction (an equity pedagogy) with her ethnically and racially diverse students in the "lowest track"…

  2. Toward an Affinity Space Methodology: Considerations for Literacy Research

    ERIC Educational Resources Information Center

    Lammers, Jayne C.; Curwood, Jen Scott; Magnifico, Alecia Marie

    2012-01-01

    As researchers seek to make sense of young people's online literacy practices and participation, questions of methodology are important to consider. In our work to understand the culture of physical, virtual and blended spheres that adolescents inhabit, we find it necessary to expand Gee's (2004) notion of affinity spaces. In this article, we draw…

  3. Peptides@mica: from affinity to adhesion mechanism.

    PubMed

    Gladytz, A; John, T; Gladytz, T; Hassert, R; Pagel, M; Risselada, H J; Naumov, S; Beck-Sickinger, A G; Abel, B

    2016-09-14

    Investigating the adsorption of peptides on inorganic surfaces, on the molecular level, is fundamental for medicinal and analytical applications. Peptides can be potent as linkers between surfaces and living cells in biochips or in implantation medicine. Here, we studied the adsorption process of the positively charged pentapeptide RTHRK, a recently identified binding sequence for surface oxidized silicon, and novel analogues thereof to negatively charged mica surfaces. Homogeneous formation of monolayers in the nano- and low micromolar peptide concentration range was observed. We propose an alternative and efficient method to both quantify binding affinity and follow adhesion behavior. This method makes use of the thermodynamic relationship between surface coverage, measured by atomic force microscopy (AFM), and the concomitant free energy of adhesion. A knowledge-based fit to the autocorrelation of the AFM images was used to correct for a biased surface coverage introduced by the finite lateral resolution of the AFM. Binding affinities and mechanisms were further explored by large scale molecular dynamics (MD) simulations. The combination of well validated MD simulations with topological data from AFM revealed a better understanding of peptide adsorption processes on the atomistic scale. We demonstrate that binding affinity is strongly determined by a peptide's ability to form salt bridges and hydrogen bonds with the surface lattice. Consequently, differences in hydrogen bond formation lead to substantial differences in binding affinity despite conservation of the peptide's overall charge. Further, MD simulations give access to relative changes in binding energy of peptide variations in comparison to a lead compound. PMID:27491508

  4. Accurate Evaluation Method of Molecular Binding Affinity from Fluctuation Frequency

    NASA Astrophysics Data System (ADS)

    Hoshino, Tyuji; Iwamoto, Koji; Ode, Hirotaka; Ohdomari, Iwao

    2008-05-01

    Exact estimation of the molecular binding affinity is significantly important for drug discovery. The energy calculation is a direct method to compute the strength of the interaction between two molecules. This energetic approach is, however, not accurate enough to evaluate a slight difference in binding affinity when distinguishing a prospective substance from dozens of candidates for medicine. Hence more accurate estimation of drug efficacy in a computer is currently demanded. Previously we proposed a concept of estimating molecular binding affinity, focusing on the fluctuation at an interface between two molecules. The aim of this paper is to demonstrate the compatibility between the proposed computational technique and experimental measurements, through several examples for computer simulations of an association of human immunodeficiency virus type-1 (HIV-1) protease and its inhibitor (an example for a drug-enzyme binding), a complexation of an antigen and its antibody (an example for a protein-protein binding), and a combination of estrogen receptor and its ligand chemicals (an example for a ligand-receptor binding). The proposed affinity estimation has proven to be a promising technique in the advanced stage of the discovery and the design of drugs.

  5. Student Engagement and Neoliberalism: Mapping an Elective Affinity

    ERIC Educational Resources Information Center

    Zepke, Nick

    2015-01-01

    The purpose of this article is to argue that student engagement, an important area for research about learning and teaching in formal higher education, has an elective affinity with neoliberalism, a hegemonic ideology in many countries of the developed world. The paper first surveys an extensive research literature examining student engagement and…

  6. Kinetic Studies of Biological Interactions By Affinity Chromatography

    PubMed Central

    Schiel, John E.; Hage, David S.

    2009-01-01

    The rates at which biological interactions occur can provide important information on the mechanism and behavior of such processes in living systems. This review will discuss how affinity chromatography can be used as a tool to examine the kinetics of biological interactions. This approach, referred to here as biointeraction chromatography, uses a column with an immobilized binding agent to examine the association or dissociation of this agent with other compounds. The use of HPLC-based affinity columns in kinetic studies has received particular attention in recent years. Advantages of using HPLC with affinity chromatography for this purpose include the ability to reuse the same ligand within a column for a large number of experiments, and the good precision and accuracy of this approach. A number of techniques are available for kinetic studies through the use of affinity columns and biointeraction chromatography. These approaches include plate height measurements, peak profiling, peak fitting, split-peak measurements, and peak decay analysis. The general principles for each of these methods are discussed in this review and some recent applications of these techniques are presented. The advantages and potential limitations of each approach are also considered. PMID:19391173

  7. Bimolecular affinity purification: a variation of TAP with multiple applications.

    PubMed

    Starokadomskyy, Petro; Burstein, Ezra

    2014-01-01

    The identification of true interacting partners of any given bait can be plagued by the nonspecific purification of irrelevant proteins. To avoid this problem, Tandem Affinity Purification (TAP) is a widely used procedure in molecular biology as this reduces the chance of nonspecific proteins being present in the final preparation. In this approach, two different affinity tags are fused to the protein bait. Herein, we review in detail a variation on the TAP procedure that we have previously developed, where the affinity moieties are placed on two different proteins that form a complex in vivo. This variation, which we refer to as Bimolecular Affinity Purification (BAP), is suited for the identification of specific molecular complexes marked by the presence of two known proteins. We have utilized BAP for characterization of molecular complexes and evaluation of proteins interaction. Another application of BAP is the isolation of ubiquitin-like proteins (UBL)-modified fractions of a given protein and characterization of the lysine-acceptor site and structure of UBL-chains. PMID:24943324

  8. Harmonic fusion and pitch affinity: Is there a direct link?

    PubMed

    Bonnard, Damien; Dauman, René; Semal, Catherine; Demany, Laurent

    2016-03-01

    Simultaneous pure tones approximately one octave apart tend to be fused perceptually and to evoke a single pitch sensation. Besides, sequentially presented pure tones show a subjective "affinity" or similarity in pitch when their frequency ratio is close to one octave. The aim of the study reported here was to determine if these two perceptual phenomena are directly related. Each stimulus was a triplet of simultaneous or successive pure tones forming frequency ratios varying across stimuli between 0.96 and 1.04 octaves. The tones were presented at a low sensation level (15 dB) within broadband threshold-equalizing noise, in order to prevent them from interacting in the cochlea when they were simultaneous. A large set of stimulus comparisons made by 18 listeners indicated that: (1) when the tones were simultaneous, maximal fusion was obtained for a mean frequency ratio deviating by less than 0.2% from one octave, and fusion decreased less rapidly above this frequency ratio than below it; (2) when the tones were presented successively, maximal pitch affinity was obtained for a mean frequency ratio significantly larger than one octave, and pitch affinity decreased more rapidly above this frequency ratio than below it. The differences between the results obtained for simultaneous and successive tones suggest that harmonic fusion and pitch affinity are unrelated phenomena. PMID:26341475

  9. RELATIVE BINDING AFFINITY OF ALKYLPHENOLS TO RAINBOW TROUT ESTROGEN RECEPTOR

    EPA Science Inventory

    RELATIVE BINDING AFFINITY OF ALKYLPHENOLS TO RAINBOW TROUT ESTROGEN RECEPTOR. T R Henry1, J S Denny2 and P K Schmieder2. USEPA, ORD, NHEERL, 1Experimental Toxicology Division and 2Mid-Continent Ecology Division, Duluth, MN, USA.
    The USEPA has been mandated to screen industria...

  10. Learning a peptide-protein binding affinity predictor with kernel ridge regression

    PubMed Central

    2013-01-01

    Background The cellular function of a vast majority of proteins is performed through physical interactions with other biomolecules, which, most of the time, are other proteins. Peptides represent templates of choice for mimicking a secondary structure in order to modulate protein-protein interaction. They are thus an interesting class of therapeutics since they also display strong activity, high selectivity, low toxicity and few drug-drug interactions. Furthermore, predicting peptides that would bind to a specific MHC alleles would be of tremendous benefit to improve vaccine based therapy and possibly generate antibodies with greater affinity. Modern computational methods have the potential to accelerate and lower the cost of drug and vaccine discovery by selecting potential compounds for testing in silico prior to biological validation. Results We propose a specialized string kernel for small bio-molecules, peptides and pseudo-sequences of binding interfaces. The kernel incorporates physico-chemical properties of amino acids and elegantly generalizes eight kernels, comprised of the Oligo, the Weighted Degree, the Blended Spectrum, and the Radial Basis Function. We provide a low complexity dynamic programming algorithm for the exact computation of the kernel and a linear time algorithm for it’s approximation. Combined with kernel ridge regression and SupCK, a novel binding pocket kernel, the proposed kernel yields biologically relevant and good prediction accuracy on the PepX database. For the first time, a machine learning predictor is capable of predicting the binding affinity of any peptide to any protein with reasonable accuracy. The method was also applied to both single-target and pan-specific Major Histocompatibility Complex class II benchmark datasets and three Quantitative Structure Affinity Model benchmark datasets. Conclusion On all benchmarks, our method significantly (p-value ≤ 0.057) outperforms the current state-of-the-art methods at predicting

  11. Myoglobin oxygen affinity in aquatic and terrestrial birds and mammals.

    PubMed

    Wright, Traver J; Davis, Randall W

    2015-07-01

    Myoglobin (Mb) is an oxygen binding protein found in vertebrate skeletal muscle, where it facilitates intracellular transport and storage of oxygen. This protein has evolved to suit unique physiological needs in the muscle of diving vertebrates that express Mb at much greater concentrations than their terrestrial counterparts. In this study, we characterized Mb oxygen affinity (P50) from 25 species of aquatic and terrestrial birds and mammals. Among diving species, we tested for correlations between Mb P50 and routine dive duration. Across all species examined, Mb P50 ranged from 2.40 to 4.85 mmHg. The mean P50 of Mb from terrestrial ungulates was 3.72±0.15 mmHg (range 3.70-3.74 mmHg). The P50 of cetaceans was similar to terrestrial ungulates ranging from 3.54 to 3.82 mmHg, with the exception of the melon-headed whale, which had a significantly higher P50 of 4.85 mmHg. Among pinnipeds, the P50 ranged from 3.23 to 3.81 mmHg and showed a trend for higher oxygen affinity in species with longer dive durations. Among diving birds, the P50 ranged from 2.40 to 3.36 mmHg and also showed a trend of higher affinities in species with longer dive durations. In pinnipeds and birds, low Mb P50 was associated with species whose muscles are metabolically active under hypoxic conditions associated with aerobic dives. Given the broad range of potential globin oxygen affinities, Mb P50 from diverse vertebrate species appears constrained within a relatively narrow range. High Mb oxygen affinity within this range may be adaptive for some vertebrates that make prolonged dives. PMID:25987728

  12. A new method of synthesizing biopolymeric affinity ligands.

    PubMed

    Chaga, G S; Guzman, R; Porath, J O

    1997-08-01

    (1) A new concept for producing soluble polymeric affinity ligands is proposed and exemplified. By solid-phase synthesis, an insoluble hydrophilic polymer is converted into an affinity gel. The gel is hydrolytically degraded to water-soluble affinity polymeric ligands which are recovered and purified. (2) A water-soluble biopolymeric metal-affinity carrier based on an iminodiacetic acid (IDA) derivative of dextran has been synthesized through the modification of Sephadex G-200 by IDA, followed by hydrolysis with dextranase and size-exclusion-chromatographic purification of the high-molecular-mass fragments. (3) The molecular size of the soluble products as a function of hydrolysis time with dextranase from Penicillium sp. was determined. The range of molecular size of the biopolymeric chelating ligand varies from around 200 Da to greater than 580 kDa. (4) The influence of three metal ions chelated with the Sephadex derivative on the hydrolysis rate and the molecular-size distribution of end products was studied. Eu3+ was found to improve the rate of solubilization. Ni2+ and Cu2+ decreased the hydrolysis rate, as compared with that of the metal-free IDA-Sephadex. (5) The method introduced here has the potential of being developed and applied as a general technology for synthesis of soluble multifunctional affinity ligands. Such ligands should be useful for liquid-phase extraction as well as for the synthesis of adsorbents with localized multiple binding sites. Other possible fields of applications are to be found in medicine, where they could be used for slow drug delivery or detoxification, and in analytical chemistry, where they could be used in various assays. PMID:9261997

  13. Novel trends in affinity biosensors: current challenges and perspectives

    NASA Astrophysics Data System (ADS)

    Arugula, Mary A.; Simonian, Aleksandr

    2014-03-01

    Molecular biorecognition processes facilitate physical and biochemical interactions between molecules in all crucial metabolic pathways. Perhaps the target analyte and the biorecognition element interactions have the most impactful use in biosensing applications. Traditional analytical sensing systems offer excellent biorecognition elements with the ability to detect and determine the presence of analytes. High affinity antibodies and DNA play an important role in the development of affinity biosensors based on electrochemical, optical and mass sensitive approaches. Advancements in this area routinely employ labels, label free, nanoparticles, multifunctional matrices, carbon nanotubes and other methods to meet the requirements of its own application. However, despite increasing affinity ceilings for conventional biosensors, the field draws back in meeting specifically important demands, such as long-term stability, ultrasensitivity, rapid detection, extreme selectivity, strong biological base, calibration, in vivo measurements, regeneration, satisfactory performance and ease of production. Nevertheless, recent efforts through this line have produced novel high-tech nanosensing systems such as ‘aptamers’ and ‘phages’ which exhibit high-throughput sensing. Aptamers and phages are powerful tools that excel over antibodies in sensibility, stability, multi-detection, in vivo measurements and regeneration. Phages are superior in stability, screening for affinity-based target molecules ranging from small to proteins and even cells, and easy production. In this review, we focus mainly on recent developments in affinity-based biosensors such as immunosensors, DNA sensors, emphasizing aptasensors and phage-based biosensors basing on novel electrochemical, optical and mass sensitive detection techniques. We also address enzyme inhibition-based biosensors and the current problems associated with the above sensors and their future perspectives.

  14. Tuning Hydrophobicity in Abiotic Affinity Reagents: Polymer Hydrogel Affinity Reagents for Molecules with Lipid-like Domains.

    PubMed

    Chou, Beverly; Mirau, Peter; Jiang, Tian; Wang, Szu-Wen; Shea, Kenneth J

    2016-05-01

    Hydrophobic interactions often dominate the associative forces between biomacromolecules. A synthetic affinity reagent must be able to exploit and optimize these interactions. We describe synthesis of abiotic affinity reagents that sequester biomacromolecules with lipid-like domains. NIPAm-based copolymer nanoparticles (NPs) containing C4-C8 hydrophobic groups were evaluated for their affinity for lipopolysaccharides (LPS), the lipophilic component of the outer membrane of Gram-negative bacteria. Optimal affinity was found for NPs incorporating a linear C4 hydrocarbon group. 1D and 2D (1)H NMR studies revealed that in water, the longer chain (C6 and C8) alkyl groups in the hydrogel NPs were engaged in intrachain association, rendering them less available to interact with LPS. Optimal LPS-NP interaction requires maximizing hydrophobicity, while avoiding side chain aggregation. Polymer compositions with high LPS binding were grafted onto agarose beads and evaluated for LPS clearance from solution; samples containing linear C4 groups also showed the highest LPS clearance capacity. PMID:27064286

  15. Engineering a reversible, high-affinity system for efficient protein purification based on the cohesin-dockerin interaction.

    PubMed

    Karpol, Alon; Kantorovich, Lia; Demishtein, Alik; Barak, Yoav; Morag, Ely; Lamed, Raphael; Bayer, Edward A

    2009-01-01

    Efficient degradation of cellulose by the anaerobic thermophilic bacterium, Clostridium thermocellum, is carried out by the multi-enzyme cellulosome complex. The enzymes on the complex are attached in a calcium-dependent manner via their dockerin (Doc) module to a cohesin (Coh) module of the cellulosomal scaffoldin subunit. In this study, we have optimized the Coh-Doc interaction for the purpose of protein affinity purification. A C. thermocellum Coh module was thus fused to a carbohydrate-binding module, and the resultant fusion protein was applied directly onto beaded cellulose, thereby serving as a non-covalent "activation" procedure. A complementary Doc module was then fused to a model protein target: xylanase T-6 from Geobacillus stearothermophilus. However, the binding to the immobilized Coh was only partially reversible upon treatment with EDTA, and only negligible amounts of the target protein were eluted from the affinity column. In order to improve protein elution, a series of truncated Docs were designed in which the calcium-coordinating function was impaired without appreciably affecting high-affinity binding to Coh. A shortened Doc of only 48 residues was sufficient to function as an effective affinity tag, and highly purified target protein was achieved directly from crude cell extracts in a single step with near-quantitative recovery of the target protein. Effective EDTA-mediated elution of the sequestered protein from the column was the key step of the procedure. The affinity column was reusable and maintained very high levels of capacity upon repeated rounds of loading and elution. Reusable Coh-Doc affinity columns thus provide an efficient and attractive approach for purifying proteins in high yield by modifying the calcium-binding loop of the Doc module. PMID:18979459

  16. Selection of a high-affinity and in vivo bioactive ssDNA aptamer against angiotensin II peptide.

    PubMed

    Heiat, Mohammad; Ranjbar, Reza; Latifi, Ali Mohammad; Rasaee, Mohammad Javad

    2016-08-01

    Unique features of aptamers have attracted interests for a broad range of applications. Aptamers are able to specifically bind to targets and inhibit their functions. This study, aimed to isolate the high affinity ssDNA aptamers against bio-regulator peptide angiotensin II (Ang II) and investigate their bioactivity in cellular and animal models. To isolate ssDNA aptamers, 12 rounds of affinity chromatography SELEX (Systematic Evolution of Ligands by EXponential enrichment) procedure were carried out. The SPR (surface plasmon resonance) and ELONA (enzyme linked oligonucleotide assay) analysis were used to determine the affinity and specificity of aptamers. The ability of selected aptamers to inhibit the proliferative effect of Ang II on human aortic vascular smooth muscle cells (HA-VSMCs) and their performance on Wistar rat urinary system and serum electrolyte levels were investigated. Two full-length aptamers (FLC112 and FLC125) with high affinity of respectively 7.52±2.44E-10 and 5.87±1.3E-9M were isolated against Ang II. The core regions of these aptamers (CRC112 and CRC125) also showed affinity of 5.33±1.15E-9 and 4.11±1.09E-9M. In vitro analysis revealed that FLC112 and FLC125 can inhibit the proliferative effect of Ang II on HA-VSMCs (P<0.05). They also significantly reduced the serum sodium level and increased the urine volume (P<0.05). The core regions of aptamers did not show high inhibitory potential against Ang II. It can be a spotlight that ssDNA aptamers have high potential for blocking Ang II. In conclusion, it appears that the researches focusing on high affinity and bioactive aptamers may lead to excellent results in blocking Ang II activity. PMID:27298205

  17. PDZ Affinity Chromatography: A general method for affinity purification of proteins based on PDZ domains and their ligands

    PubMed Central

    Walkup, Ward G.; Kennedy, Mary B.

    2014-01-01

    PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ~ 90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized that the specific, strong interactions of PDZ domains with their ligands would make them well suited for use in affinity chromatography. Here we describe a novel affinity chromatography method applicable for the purification of proteins that contain PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We created a series of affinity resins comprised of PDZ domains from the scaffold protein PSD-95, or from neuronal nitric oxide synthase (nNOS), coupled to solid supports. We used them to purify heterologously expressed neuronal proteins or protein domains containing endogenous PDZ domain ligands, eluting the proteins with free PDZ domain peptide ligands. We show that Proteins of Interest (POIs) lacking endogenous PDZ domain ligands can be engineered as fusion products containing C-terminal PDZ domain ligand peptides or internal, N- or C-terminal PDZ domains and then can be purified by the same method. Using this method, we recovered recombinant GFP fused to a PDZ-domain ligand in active form as verified by fluorescence yield. Similarly, chloramphenicol acetyltransferase (CAT) and β-Galactosidase (LacZ) fused to a C-terminal PDZ domain ligand or an N-terminal PDZ domain were purified in active form as assessed by enzymatic assay. In general, PDZ domains and ligands derived from PSD-95 were superior to those from nNOS for this method. PDZ Domain Affinity Chromatography promises to be a versatile and effective method for purification of a wide variety of natural and recombinant proteins. PMID:24607360

  18. Structural Basis of Species-Dependent Differential Affinity of 6-Alkoxy-5-Aryl-3-Pyridinecarboxamide Cannabinoid-1 Receptor Antagonists

    PubMed Central

    Iyer, Malliga R.; Cinar, Resat; Liu, Jie; Godlewski, Grzegorz; Szanda, Gergö; Puhl, Henry; Ikeda, Stephen R.; Deschamps, Jeffrey; Lee, Yong-Sok; Steinbach, Peter J.

    2015-01-01

    6-Alkoxy-5-aryl-3-pyridincarboxamides, including the brain-penetrant compound 14g [5-(4-chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxy-cyclohexyl]-3-pyridinecarboxamide] and its peripherally restricted analog 14h [5-(4-chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2-methoxyethoxy)-3-pyridinecarboxamide], have been recently introduced as selective, high-affinity antagonists of the human cannabinoid-1 receptor (hCB1R). Binding analyses revealed two orders of magnitude lower affinity of these compounds for mouse and rat versus human CB1R, whereas the affinity of rimonabant is comparable for all three CB1Rs. Modeling of ligand binding to CB1R and binding assays with native and mutant (Ile105Met) hCB1Rs indicate that the Ile105 to Met mutation in rodent CB1Rs accounts for the species-dependent affinity of 14g and 14h. Our work identifies Ile105 as a new pharmacophore component for developing better hCB1R antagonists and invalidates rodent models for assessing the antiobesity efficacy of 14g and 14h. PMID:26013543

  19. Evaluation of Taylor dispersion injections: determining kinetic/affinity interaction constants and diffusion coefficients in label-free biosensing.

    PubMed

    Quinn, John G

    2012-02-15

    In label-free biomolecular interaction analysis, a standard injection provides an injection of uniform analyte concentration. An alternative approach exploiting Taylor dispersion produces a continuous analyte titration allowing a full analyte dose response to be recorded in a single injection. The enhanced biophysical characterization that is possible with this new technique is demonstrated using a commercially available surface plasmon resonance-based biosensor. A kinetic interaction model was fitted locally to Taylor dispersion curves for estimation of the analyte diffusion coefficient in addition to affinity/kinetic constants. Statistical confidence in the measured parameters from a single Taylor dispersion injection was comparable to that obtained for global analysis of multiple standard injections. The affinity constants for multisite interactions were resolved with acceptable confidence limits. Importantly, a single analyte injection could be treated as a high-resolution real-time affinity isotherm and was demonstrated using the complex two-site interaction of warfarin with human serum albumin. In all three model interactions tested, the kinetic/affinity constants compared favorably with those obtained from standard kinetic analysis and the estimates of analyte diffusion coefficients were in good agreement with the expected values. PMID:22197422

  20. Coupling Isotachophoresis with Affinity Chromatography for Rapid and Selective Purification with High Column Utilization, Part 1: Theory

    PubMed Central

    2015-01-01

    We present a novel technique that couples isotachophoresis (ITP) with affinity chromatography (AC) to achieve rapid, selective purification with high column utilization. ITP simultaneously preconcentrates an analyte and purifies it, based on differences in mobility of sample components, excluding species that may foul or compete with the target at the affinity substrate. ITP preconcentration accelerates the affinity reaction, reducing assay time, improving column utilization, and allowing for capture of targets with higher dissociation constants. Furthermore, ITP-AC separates the target and contaminants into nondiffusing zones, thus achieving high resolution in a short distance and time. We present an analytical model for spatiotemporal dynamics of ITP-AC. We identify and explore the effect of key process parameters, including target distribution width and height, ITP zone velocity, forward and reverse reaction constants, and probe concentration on necessary affinity region length, assay time, and capture efficiency. Our analytical approach shows collapse of these variables to three nondimensional parameters. The analysis yields simple analytical relations for capture length and capture time in relevant ITP-AC regimes, and it demonstrates how ITP greatly reduces assay time and improves column utilization. In the second part of this two-part series, we will present experimental validation of our model and demonstrate ITP-AC separation of the target from 10,000-fold more-abundant contaminants. PMID:24937679

  1. Affinity learning with diffusion on tensor product graph.

    PubMed

    Yang, Xingwei; Prasad, Lakshman; Latecki, Longin Jan

    2013-01-01

    In many applications, we are given a finite set of data points sampled from a data manifold and represented as a graph with edge weights determined by pairwise similarities of the samples. Often the pairwise similarities (which are also called affinities) are unreliable due to noise or due to intrinsic difficulties in estimating similarity values of the samples. As observed in several recent approaches, more reliable similarities can be obtained if the original similarities are diffused in the context of other data points, where the context of each point is a set of points most similar to it. Compared to the existing methods, our approach differs in two main aspects. First, instead of diffusing the similarity information on the original graph, we propose to utilize the tensor product graph (TPG) obtained by the tensor product of the original graph with itself. Since TPG takes into account higher order information, it is not a surprise that we obtain more reliable similarities. However, it comes at the price of higher order computational complexity and storage requirement. The key contribution of the proposed approach is that the information propagation on TPG can be computed with the same computational complexity and the same amount of storage as the propagation on the original graph. We prove that a graph diffusion process on TPG is equivalent to a novel iterative algorithm on the original graph, which is guaranteed to converge. After its convergence we obtain new edge weights that can be interpreted as new, learned affinities. We stress that the affinities are learned in an unsupervised setting. We illustrate the benefits of the proposed approach for data manifolds composed of shapes, images, and image patches on two very different tasks of image retrieval and image segmentation. With learned affinities, we achieve the bull's eye retrieval score of 99.99 percent on the MPEG-7 shape dataset, which is much higher than the state-of-the-art algorithms. When the data

  2. Affinity ligands for glycoprotein purification based on the multi-component Ugi reaction.

    PubMed

    Chen, Chen; Khoury, Graziella El; Lowe, Christopher R

    2014-10-15

    One challenge facing the purification of therapeutic glycoproteins by affinity chromatography is creating ligands specific for the glycan moiety. Affinity chromatography of glycoproteins is currently conducted with immobilized lectins or boronates, although biomimetic ligands could present a more desirable option. This work describes the rational design and combinatorial synthesis of carbohydrate-binding ligands based on the solid phase multi-component Ugi reaction. An aldehyde-functionalized Sepharose™ solid support constitutes one component (aldehyde) in the four-component reaction, while the other three components (a primary/secondary amine, a carboxylic acid and an isocyanide) are varied in a combinatorial fashion to generate a tri-substituted Ugi scaffold which provides a degree of rigidity and is functionally suitable for interacting with the glycan moiety of glycoproteins. An Ugi library containing 48 ligands was initially screened against glucose oxidase (GOx) as the model glycoprotein to identify a candidate ligand, A13C24I8, which showed affinity to GOx through its carbohydrate moiety. Immobilized ligand A13C24I8 demonstrated a static binding capacity of 16.7mg GOx/ml resin and an apparent dissociation constant (Kd) of 1.45×10(-6)M at pH 7.4. The adsorbent can also bind 8.1mg AGP/ml resin and displays an apparent affinity constant Kd=1.44×10(-5)M. The ligand has a sugar specificity in the following sequence: sorbitol>fructose>mannitol>ribose>arabinose>xylose>galactose>mannose>glucose>fructose; however, it did not display any specificity for sialic acid or methyl α-D-glycosides. A control ligand, generated by substitution of C24 (3-carboxyphenylboronic acid) with C7 (4-hydroxyphenyl acetic acid), failed to show affinity to the carbohydrate moiety, supporting the importance of the role that boronic acid group plays in sugar binding. GOx spiked E. coli samples were loaded onto immobilized ligand A13C24I8, 3-aminophenylboronic acid (APBA) and

  3. Differentiation of extrastriatal dopamine D2 receptor density and affinity in the human brain using PET.

    PubMed

    Olsson, Hans; Halldin, Christer; Farde, Lars

    2004-06-01

    Dopaminergic neurotransmission in extrastriatal regions may play a crucial role in the pathophysiology and treatment of neuropsychiatric disorders. The high-affinity radioligands [(11)C]FLB 457, [(123)I]epidepride, and [(18)F]fallypride are now used in clinical studies to measure these low-density receptor populations in vivo. However, a single determination of the regional binding potential (BP) does not differentiate receptor density (B(max)) from the apparent affinity (K(D)). In this positron emission tomography (PET) study, we measured extrastriatal dopamine D2 receptor density (B(max)) and apparent affinity (K(D)) in 10 healthy subjects using an in vivo saturation approach. Each subject participated in two to three PET measurements with different specific radioactivity of [(11)C]FLB 457. The commonly used simplified reference tissue model (SRTM) was used in a comparison of BP values with the B(max) values obtained from the saturation analysis. The calculated regional receptor density values were of the same magnitude (0.33-1.68 nM) and showed the same rank order as reported from postmortem studies, that is, in descending order thalamus, lateral temporal cortex, anterior cinguli, and frontal cortex. The affinity ranged from 0.27 to 0.43 nM, that is, approximately 10-20 times the value found in vitro (20 pM). The area under the cerebellar time activity curve (TAC) was slightly lower (11 +/- 8%, mean +/- SD, P = 0.004, n = 10) after injection of low as compared with high specific radioactivity, indicating sensitivity to the minute density of dopamine D2 receptors in the this region. The results of the present study support that dopamine D2 receptor density and affinity can be differentiated in low-density regions using a saturation approach. There was a significant (P < 0.001) correlation between the binding potential calculated with SRTM and the receptor density (B(max)), which supports the use of BP in clinical studies where differentiation of B(max) and K

  4. Affinity and specificity of interactions between Nedd4 isoforms and the epithelial Na+ channel.

    PubMed

    Henry, Pauline C; Kanelis, Voula; O'Brien, M Christine; Kim, Brian; Gautschi, Ivan; Forman-Kay, Julie; Schild, Laurent; Rotin, Daniela

    2003-05-30

    The epithelial Na+ channel (alphabetagammaENaC) regulates salt and fluid homeostasis and blood pressure. Each ENaC subunit contains a PY motif (PPXY) that binds to the WW domains of Nedd4, a Hect family ubiquitin ligase containing 3-4 WW domains and usually a C2 domain. It has been proposed that Nedd4-2, but not Nedd4-1, isoforms can bind to and suppress ENaC activity. Here we challenge this notion and show that, instead, the presence of a unique WW domain (WW3*) in either Nedd4-2 or Nedd4-1 determines high affinity interactions and the ability to suppress ENaC. WW3* from either Nedd4-2 or Nedd4-1 binds ENaC-PY motifs equally well (e.g. Kd approximately 10 microm for alpha- or betaENaC, 3-6-fold higher affinity than WW4), as determined by intrinsic tryptophan fluorescence. Moreover, dNedd4-1, which naturally contains a WW3* instead of WW2, is able to suppress ENaC function equally well as Nedd4-2. Homology models of the WW3*.betaENaC-PY complex revealed that a Pro and Ala conserved in all WW3*, but not other Nedd4-WW domains, help form the binding pocket for PY motif prolines. Extensive contacts are formed between the betaENaC-PY motif and the Pro in WW3*, and the small Ala creates a large pocket to accommodate the peptide. Indeed, mutating the conserved Pro and Ala in WW3* reduces binding affinity 2-3-fold. Additionally, we demonstrate that mutations in PY motif residues that form contacts with the WW domain based on our previously solved structure either abolish or severely reduce binding affinity to the WW domain and that the extent of binding correlates with the level of ENaC suppression. Independently, we show that a peptide encompassing the PY motif of sgk1, previously proposed to bind to Nedd4-2 and alter its ability to regulate ENaC, does not bind (or binds poorly) the WW domains of Nedd4-2. Collectively, these results suggest that high affinity of WW domain-PY-motif interactions rather than affiliation with Nedd4-1/Nedd-2 is critical for ENaC suppression

  5. Influence of Sulfolane on ESI-MS Measurements of Protein-Ligand Affinities

    NASA Astrophysics Data System (ADS)

    Yao, Yuyu; Richards, Michele R.; Kitova, Elena N.; Klassen, John S.

    2016-03-01

    The results of an investigation into the influence of sulfolane, a commonly used supercharging agent, on electrospray ionization mass spectrometry (ESI-MS) measurements of protein-ligand affinities are described. Binding measurements carried out on four protein-carbohydrate complexes, lysozyme with β- d-GlcNAc-(1→4)-β- d-GlcNAc-(1→4)-β- d-GlcNAc-(1→4)- d-GlcNAc, a single chain variable fragment and α- d-Gal-(1→2)-[α- d-Abe-(1→3)]-α- d-Man-OCH3, cholera toxin B subunit homopentamer with β- d-Gal-(1→3)-β- d-GalNAc-(1→4)[α- d-Neu5Ac-(2→3)]-β- d-Gal-(1→4)-β- d-Glc, and a fragment of galectin 3 and α- l-Fuc-(1→2)-β- d-Gal-(1→3)-β- d-GlcNAc-(1→3)-β- d-Gal-(1→4)-β- d-Glc, revealed that sulfolane generally reduces the apparent (as measured by ESI-MS) protein-ligand affinities. To establish the origin of this effect, a detailed study was undertaken using the lysozyme-tetrasaccharide interaction as a model system. Measurements carried out using isothermal titration calorimetry (ITC), circular dichroism, and nuclear magnetic resonance spectroscopies reveal that sulfolane reduces the binding affinity in solution but does not cause any significant change in the higher order structure of lysozyme or to the intermolecular interactions. These observations confirm that changes to the structure of lysozyme in bulk solution are not responsible for the supercharging effect induced by sulfolane. Moreover, the agreement between the ESI-MS and ITC-derived affinities indicates that there is no dissociation of the complex during ESI or in the gas phase (i.e., in-source dissociation). This finding suggests that supercharging of lysozyme by sulfolane is not related to protein unfolding during the ESI process. Binding measurements performed using liquid sample desorption ESI-MS revealed that protein supercharging with sulfolane can be achieved without a reduction in affinity.

  6. Gini covariance matrix and its affine equivariant version

    NASA Astrophysics Data System (ADS)

    Weatherall, Lauren Anne

    Gini's mean difference (GMD) and its derivatives such as Gini index have been widely used as alternative measures of variability over one century in many research fields especially in finance, economics and social welfare. In this dissertation, we generalize the univariate GMD to the multivariate case and propose a new covariance matrix so called the Gini covariance matrix (GCM). The extension is natural, which is based on the covariance representation of GMD with the notion of multivariate spatial rank function. In order to gain the affine equivariance property for GCM, we utilize the transformation-retransformation (TR) technique and obtain TR version GCM that turns out to be a symmetrized M-functional. Indeed, both GCMs are symmetrized approaches based on the difference of two independent variables without reference of a location, hence avoiding some arbitrary definition of location for non-symmetric distributions. We study the properties of both GCMs. They possess the so-called independence property, which is highly important, for example, in independent component analysis. Influence functions of two GCMs are derived to assess their robustness. They are found to be more robust than the regular covariance matrix but less robust than Tyler and Dumbgen M-functional. Under elliptical distributions, the relationship between the scatter parameter and the two GCM are obtained. With this relationship, principal component analysis (PCA) based on GCM is possible. Estimation of two GCMs is presented. We study asymptotical behavior of the estimators. √n-consistency and asymptotical normality of estimators are established. Asymptotic relative efficiency (ARE) of TR-GCM estimator with respect to sample covariance matrix is compared to that of Tyler and Dumbgen M-estimators. With little loss on efficiency (< 2%) in the normal case, it gains high efficiency for heavy-tailed distributions. Finite sample behavior of Gini estimators is explored under various models using two

  7. Self-affine nature of the stress-strain behavior of thin fiber networks

    NASA Astrophysics Data System (ADS)

    Balankin, Alexander S.; Susarrey, Orlando; Bravo, Armando

    2001-12-01

    The stress-strain behavior of toilet paper is studied. We find that the damaged parts of stress-strain curves possess a self-affine scaling invariance. Moreover, we find that the stress-strain behavior and the rupture line roughness are characterized by the same scaling (Hurst) exponent H, which is not universal: rather it changes from sample to sample. The variations on H are mainly due to fluctuations in the paper structure, which are larger than statistical errors within a sample. Furthermore, the same exponent governs the changes in the stress-strain curve as the strain rate increases. The fractal damage model is employed to explain experimental observations.

  8. Einstein-aether theory, violation of Lorentz invariance, and metric-affine gravity

    SciTech Connect

    Heinicke, Christian; Baekler, Peter; Hehl, Friedrich W.

    2005-07-15

    We show that the Einstein-aether theory of Jacobson and Mattingly (J and M) can be understood in the framework of the metric-affine (gauge theory of) gravity (MAG). We achieve this by relating the aether vector field of J and M to certain post-Riemannian nonmetricity pieces contained in an independent linear connection of spacetime. Then, for the aether, a corresponding geometrical curvature-square Lagrangian with a massive piece can be formulated straightforwardly. We find an exact spherically symmetric solution of our model.

  9. Linear Interaction Energy Based Prediction of Cytochrome P450 1A2 Binding Affinities with Reliability Estimation

    PubMed Central

    Capoferri, Luigi; Verkade-Vreeker, Marlies C. A.; Buitenhuis, Danny; Commandeur, Jan N. M.; Pastor, Manuel; Vermeulen, Nico P. E.; Geerke, Daan P.

    2015-01-01

    Prediction of human Cytochrome P450 (CYP) binding affinities of small ligands, i.e., substrates and inhibitors, represents an important task for predicting drug-drug interactions. A quantitative assessment of the ligand binding affinity towards different CYPs can provide an estimate of inhibitory activity or an indication of isoforms prone to interact with the substrate of inhibitors. However, the accuracy of global quantitative models for CYP substrate binding or inhibition based on traditional molecular descriptors can be limited, because of the lack of information on the structure and flexibility of the catalytic site of CYPs. Here we describe the application of a method that combines protein-ligand docking, Molecular Dynamics (MD) simulations and Linear Interaction Energy (LIE) theory, to allow for quantitative CYP affinity prediction. Using this combined approach, a LIE model for human CYP 1A2 was developed and evaluated, based on a structurally diverse dataset for which the estimated experimental uncertainty was 3.3 kJ mol-1. For the computed CYP 1A2 binding affinities, the model showed a root mean square error (RMSE) of 4.1 kJ mol-1 and a standard error in prediction (SDEP) in cross-validation of 4.3 kJ mol-1. A novel approach that includes information on both structural ligand description and protein-ligand interaction was developed for estimating the reliability of predictions, and was able to identify compounds from an external test set with a SDEP for the predicted affinities of 4.6 kJ mol-1 (corresponding to 0.8 pKi units). PMID:26551865

  10. Comparative oxygen affinity of fish and mammalian myoglobins.

    PubMed

    Nichols, J W; Weber, L J

    1989-01-01

    Myoglobins from rat, coho salmon (Oncorhynchus kisutch), buffalo sculpin (Enophrys bison) hearts, and yellowfin tuna (Thunnus albacares) red skeletal muscle were partially purified and their O2 binding affinities determined. Commercially prepared sperm whale myoglobin was employed as an internal standard. Tested at 20 degrees C, myoglobins from salmon and sculpin bound O2 with lower affinity than myoglobins from the rat or sperm whale. Oxygen binding studies at 12 degrees C and 37 degrees C suggest that this difference is adaptive, permitting myoglobins from cold-adapted fish to function at physiologically relevant temperatures. Taken together, purification and O2 binding data obtained in this study reveal a previously unrecognized diversity of myoglobin structure and function. PMID:2760286

  11. Isotope shift in the electron affinity of lithium

    SciTech Connect

    Bubin, Sergiy; Komasa, Jacek; Stanke, Monika; Adamowicz, Ludwik

    2009-12-21

    Very accurate electron affinity (EA) calculations of {sup 6}Li and {sup 7}Li (and {sup {infinity}L}i) have been performed using explicitly correlated Gaussian functions and a variational approach that explicitly includes the nuclear motion in the calculations (i.e., the approach that does not assume the Born-Oppenheimer approximation). The leading relativistic and quantum electrodynamics corrections to the electron affinities were also calculated. The results are the most accurate theoretical values obtained for the studied systems to date. Our best estimates of the {sup 7}Li and {sup 6}Li EAs are 4984.9842(30) and 4984.9015(30) cm{sup -1}, respectively, and of the {sup 7}Li/{sup 6}Li EA isotope shift is 0.0827 cm{sup -1}.

  12. Complex high affinity interactions occur between MHCI and superantigens

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Herpich, A. R.; Spooner, B. S. (Principal Investigator)

    1998-01-01

    Staphylococcal enterotoxins A and C1 (SEA or SEC1) bound to major histocompatibility-I (MHCI) molecules with high affinity (binding constants ranging from 1.1 microM to 79 nM). SEA and SEC1 directly bound MHCI molecules that had been captured by monoclonal antibodies specific for H-2Kk, H-2Dk, or both. In addition, MHCI-specific antibodies inhibited the binding of SEC1 to LM929 cells and SEA competitively inhibited SEC1 binding; indicating that the superantigens bound to MHCI on the cell surface. The affinity and number of superantigen binding sites differed depending on whether MHCI was expressed in the membrane of LM929 cells or whether it was captured. These data support the hypothesis that MHCI molecules can serve as superantigen receptors.

  13. Affinity Propagation Clustering of Measurements for Multiple Extended Target Tracking.

    PubMed

    Zhang, Tao; Wu, Renbiao

    2015-01-01

    More measurements are generated by the target per observation interval, when the target is detected by a high resolution sensor, or there are more measurement sources on the target surface. Such a target is referred to as an extended target. The probability hypothesis density filter is considered an efficient method for tracking multiple extended targets. However, the crucial problem of how to accurately and effectively partition the measurements of multiple extended targets remains unsolved. In this paper, affinity propagation clustering is introduced into measurement partitioning for extended target tracking, and the elliptical gating technique is used to remove the clutter measurements, which makes the affinity propagation clustering capable of partitioning the measurement in a densely cluttered environment with high accuracy. The Gaussian mixture probability hypothesis density filter is implemented for multiple extended target tracking. Numerical results are presented to demonstrate the performance of the proposed algorithm, which provides improved performance, while obviously reducing the computational complexity. PMID:26370998

  14. Affinity Propagation Clustering of Measurements for Multiple Extended Target Tracking

    PubMed Central

    Zhang, Tao; Wu, Renbiao

    2015-01-01

    More measurements are generated by the target per observation interval, when the target is detected by a high resolution sensor, or there are more measurement sources on the target surface. Such a target is referred to as an extended target. The probability hypothesis density filter is considered an efficient method for tracking multiple extended targets. However, the crucial problem of how to accurately and effectively partition the measurements of multiple extended targets remains unsolved. In this paper, affinity propagation clustering is introduced into measurement partitioning for extended target tracking, and the elliptical gating technique is used to remove the clutter measurements, which makes the affinity propagation clustering capable of partitioning the measurement in a densely cluttered environment with high accuracy. The Gaussian mixture probability hypothesis density filter is implemented for multiple extended target tracking. Numerical results are presented to demonstrate the performance of the proposed algorithm, which provides improved performance, while obviously reducing the computational complexity. PMID:26370998

  15. The Weyl-Cartan Space Problem in Purely Affine Theory

    NASA Astrophysics Data System (ADS)

    von Borzeszkowski, Horst-Heino; Treder, Hans-Jürgen

    1997-04-01

    According to Poincaré, only the ``epistemological sum of geometry and physics is measurable". Of course, there are requirements of measurement to be imposed on geometry because otherwise the theory resting on this geometry cannot be physically interpreted. In particular, the Weyl--Cartan space problem must be solved, i.e., it must be guaranteed that the comparison of distances is compatible with the Levi-Civita transport. In the present paper, we discuss these requirements of measurement and show that in the (purely affine) Einstein-Schrödinger unified field theory the solution of the Weyl-Cartan space problem simultaneously determines the matter via Einstein's equations. Here the affine field $\\Gamma^ikl$ represents Poincaré's sum, and the solution of the space problem means its splitting in a metrical space and in matter fields, where the latter are given by the torsion tensor $\\Gamma^i_{[kl]}$.

  16. Affinity-Driven Immobilization of Proteins to Hematite Nanoparticles.

    PubMed

    Zare-Eelanjegh, Elaheh; Bora, Debajeet K; Rupper, Patrick; Schrantz, Krisztina; Thöny-Meyer, Linda; Maniura-Weber, Katharina; Richter, Michael; Faccio, Greta

    2016-08-10

    Functional nanoparticles are valuable materials for energy production, bioelectronics, and diagnostic devices. The combination of biomolecules with nanosized material produces a new hybrid material with properties that can exceed the ones of the single components. Hematite is a widely available material that has found application in various sectors such as in sensing and solar energy production. We report a single-step immobilization process based on affinity and achieved by genetically engineering the protein of interest to carry a hematite-binding peptide. Fabricated hematite nanoparticles were then investigated for the immobilization of the two biomolecules C-phycocyanin (CPC) and laccase from Bacillus pumilus (LACC) under mild conditions. Genetic engineering of biomolecules with a hematite-affinity peptide led to a higher extent of protein immobilization and enhanced the catalytic activity of the enzyme. PMID:27429157

  17. Affinity purification of proteins binding to GST fusion proteins.

    PubMed

    Swaffield, J C; Johnston, S A

    2001-05-01

    This unit describes the use of proteins fused to glutathione-S-transferase (GST fusion proteins) to affinity purify other proteins, a technique also known as GST pulldown purification. The describes a strategy in which a GST fusion protein is bound to agarose affinity beads and the complex is then used to assay the binding of a specific test protein that has been labeled with [35S]methionine by in vitro translation. However, this method can be adapted for use with other types of fusion proteins; for example, His6, biotin tags, or maltose-binding protein fusions (MBP), and these may offer particular advantages. A describes preparation of an E. coli extract that is added to the reaction mixture with purified test protein to reduce nonspecific binding. PMID:18265191

  18. Stable high capacity, F-actin affinity column

    SciTech Connect

    Luna, E.J.; Wang, Y.L.; Voss, E.W. Jr.; Branton, D.; Taylor, D.L.

    1982-11-10

    A high capacity F-actin affinity matrix is constructed by binding fluorescyl-actin to rabbit anti-fluorescein IgG that is covalently bound to Sepharose 4B. When stabilized with phalloidin, the actin remains associated with the Sepharose beads during repeated washes, activates the ATPase activity of myosin subfragment 1, and specifically binds /sup 125/I-heavy meromyosin and /sup 125/I-tropomyosin. The associations between the F-actin-binding proteins are monitored both by affinity chromatography and by a rapid, low speed sedimentation assay. Anti-fluorescein IgG-Sepharose should be generally useful as a matrix for the immobilization of proteins containing accessible, covalently bound fluorescein groups.

  19. Affinity chromatography based on a combinatorial strategy for rerythropoietin purification.

    PubMed

    Martínez-Ceron, María C; Marani, Mariela M; Taulés, Marta; Etcheverrigaray, Marina; Albericio, Fernando; Cascone, Osvaldo; Camperi, Silvia A

    2011-05-01

    Small peptides containing fewer than 10 amino acids are promising ligand candidates with which to build affinity chromatographic systems for industrial protein purification. The application of combinatorial peptide synthesis strategies greatly facilitates the discovery of suitable ligands for any given protein of interest. Here we sought to identify peptide ligands with affinity for recombinant human erythropoietin (rhEPO), which is used for the treatment of anemia. A combinatorial library containing the octapeptides X-X-X-Phe-X-X-Ala-Gly, where X = Ala, Asp, Glu, Phe, His, Leu, Asn, Pro, Ser, or Thr, was synthesized on HMBA-ChemMatrix resin by the divide-couple-recombine method. For the library screening, rhEPO was coupled to either Texas Red or biotin. Fluorescent beads or beads showing a positive reaction with streptavidin-peroxidase were isolated. After cleavage, peptides were sequenced by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Fifty-seven beads showed a positive reaction. Peptides showing more consensuses were synthesized, and their affinity to rhEPO was assessed using a plasma resonance biosensor. Dissociation constant values in the range of 1-18 μM were obtained. The best two peptides were immobilized on Sepharose, and the resultant chromatographic matrixes showed affinity for rhEPO with dissociation constant values between 1.8 and 2.7 μM. Chinese hamster ovary (CHO) cell culture supernatant was spiked with rhEPO, and the artificial mixture was loaded on Peptide-Sepharose columns. The rhEPO was recovered in the elution fraction with a yield of 90% and a purity of 95% and 97% for P1-Sepharose and P2-Sepharose, respectively. PMID:21495625

  20. Cohomology of Various Completions of Quasicoherent Sheaves on Affines

    PubMed Central

    Laudal, Olav Arnfinn

    1972-01-01

    Let O be a complete discrete valuation ring and let A be a commutative O-algebra. Let M be any A-module. In this paper, a class of completions M̃ on the affine X corresponding to A, which includes, e.g., the Washnitzer-Monsky completion [1], and the full completion is studied. We then prove that for all of these completions we have, Hi(X,M̃+) = O for i ≥ 1, H°(X,M̃+) = M+. PMID:16592014

  1. Selective high affinity polydentate ligands and methods of making such

    SciTech Connect

    DeNardo, Sally; DeNardo, Gerald; Balhorn, Rodney

    2010-02-16

    This invention provides novel polydentate selective high affinity ligands (SHALs) that can be used in a variety of applications in a manner analogous to the use of antibodies. SHALs typically comprise a multiplicity of ligands that each bind different region son the target molecule. The ligands are joined directly or through a linker thereby forming a polydentate moiety that typically binds the target molecule with high selectivity and avidity.

  2. Purification of muscarinic acetylcholine receptors by affinity chromatography.

    PubMed Central

    André, C; De Backer, J P; Guillet, J C; Vanderheyden, P; Vauquelin, G; Strosberg, A D

    1983-01-01

    Calf forebrain homogenates contain 2.8 pM muscarinic acetylcholine receptors per mg of protein. [3H]Antagonist saturation binding experiments under equilibrium conditions revealed a single class of sites with equilibrium dissociation constants of 0.82 nM for [3H]dexetimide and 0.095 nM for [3H]quinuclidinyl benzilate. Displacement binding studies with agonists revealed the presence of low and high affinity sites. Here we describe the solubilization of muscarinic acetylcholine receptors with digitonin and their purification by affinity chromatography using an affinity gel which consisted of dexetimide coupled to Affi-Gel 10 (i.e., carboxy N-hydroxysuccinimide esters linked via a 1 nm spacer arm to agarose beads). Purified proteins were obtained by specific elution with muscarinic drugs, i.e., the antagonist atropine and the irreversible ligand propylbenzilylcholine mustard. SDS-polyacrylamide gel electrophoresis of the radioiodinated purified preparations revealed a major 70-K protein. Images Fig. 3. PMID:6605245

  3. Crystal structures of fusion proteins with large-affinity tags.

    PubMed

    Smyth, Douglas R; Mrozkiewicz, Marek K; McGrath, William J; Listwan, Pawel; Kobe, Bostjan

    2003-07-01

    The fusion of a protein of interest to a large-affinity tag, such as the maltose-binding protein (MBP), thioredoxin (TRX), or glutathione-S-transferase (GST), can be advantageous in terms of increased expression, enhanced solubility, protection from proteolysis, improved folding, and protein purification via affinity chromatography. Unfortunately, crystal growth is hindered by the conformational heterogeneity induced by the fusion tag, requiring that the tag is removed by a potentially problematic cleavage step. The first three crystal structures of fusion proteins with large-affinity tags have been reported recently. All three structures used a novel strategy to rigidly fuse the protein of interest to MBP via a short three- to five-amino acid spacer. This strategy has the potential to aid structure determination of proteins that present particular experimental challenges and are not conducive to more conventional crystallization strategies (e.g., membrane proteins). Structural genomics initiatives may also benefit from this approach as a way to crystallize problematic proteins of significant interest. PMID:12824478

  4. Use of protein-protein interactions in affinity chromatography.

    PubMed

    Muronetz, V I; Sholukh, M; Korpela, T

    2001-10-30

    Biospecific recognition between proteins is a phenomenon that can be exploited for designing affinity-chromatographic purification systems for proteins. In principle, the approach is straightforward, and there are usually many alternative ways, since a protein can be always found which binds specifically enough to the desired protein. Routine immunoaffinity chromatography utilizes the recognition of antigenic epitopes by antibodies. However, forces involved in protein-protein interactions as well the forces keeping the three-dimensional structures of proteins intact are complicated, and proteins are easily unfolded by various factors with unpredictable results. Because of this and because of the generally high association strength between proteins, the correct adjustment of binding forces between an immobilized protein and the protein to be purified as well as the release of bound proteins in biologically active form from affinity complexes are the main problem. Affinity systems involving interactions like enzyme-enzyme, subunit-oligomer, protein-antibody, protein-chaperone and the specific features involved in each case are presented as examples. This article also aims to sketch prospects for further development of the use of protein-protein interactions for the purification of proteins. PMID:11694271

  5. An Early Cambrian problematic fossil: Vetustovermis and its possible affinities

    PubMed Central

    Chen, Jun-yuan; Huang, Di-ying; Bottjer, David J

    2005-01-01

    The Early Cambrian problematic fossil Vetustovermis (Glaessner 1979 Alcheringa 3, 21–31) was described as an annelid or arthropod. Anatomical analysis of 17 new specimens from the Lower Cambrian Maotianshan Shale at Anning, Kunming (South China) does not support its affinities with annelids or arthropods. Anatomical features instead resemble other animal groups including modern flatworms, nemertines and molluscs. The presence of a pelagic slug-like form and ventral foot, as well as a head with eyes and tentacles indicates a possible affinity with molluscs, but these characters are not present only in molluscs; some of them are shared with other animal groups, including flatworms and nemertines. For example, a ventral foot-like structure is found in nemertines, ‘turbellarians’, and some polychaete groups. The well differentiated head is seen in separate bilaterian groups, but among molluscs it did not occur before the evolutionary level of the Conchifera. Unlike the ctenia-gills in molluscs, the gills in Vetustovermis are bar-like. All the characters displayed in this 525 million-year old soft-bodied animal fail to demonstrate clear affinity with molluscs or any other known extant or extinct animal groups, but argue for representing an independently evolved animal group, which flourished in Early Cambrian and possibly in Middle Cambrian time. PMID:16191609

  6. Virtual-real spatial information visualization registration using affine representations

    NASA Astrophysics Data System (ADS)

    Wu, Xueling; Ren, Fu; Du, Qingyun

    2009-10-01

    Virtual-real registration in Outdoor Augmented Reality is committed to enhance user's spatial cognition by overlaying virtual geographical objects on real scene. According to analyze fiducial detection registration method in indoor AR, for the purpose of avoiding complex and tedious process of position tracking and camera calibration in traditional registration methods, it puts forward and practices a virtual-real spatial information visualization registration method using affine representations. Based on the observation from Koenderink and van Doorn, Ullman and Basri in 1991 which is given a set of four or more non-coplanar 3D points, the projection of all points in the set can be computed as a linear combination of the projection of just four of the points, it sets up global affine coordinate system in light of world coordinates, camera coordinates and virtual coordinates and extracts four feature points from scene image and calculates the global affine coordinates of key points of virtual objects. Then according to a linear homogeneous coordinates of the four feature point's projection, it calculates projection pixel coordinates of key points of virtual objects. In addition, it proposes an approach to obtain pixel relative depth for hidden surface removal. Finally, by a case study, it verifies the feasibility and efficiency of the registration methods. The method would not only explore a new research direction for Geographical Information Science, but also would provide location-based information and services for outdoor AR.

  7. Affitins for protein purification by affinity magnetic fishing.

    PubMed

    Fernandes, Cláudia S M; Dos Santos, Raquel; Ottengy, Stella; Viecinski, Aline Canani; Béhar, Ghislaine; Mouratou, Barbara; Pecorari, Frédéric; Roque, A Cecília A

    2016-07-29

    Currently most economical and technological bottlenecks in protein production are placed in the downstream processes. With the aim of increasing the efficiency and reducing the associated costs, various affinity ligands have been developed. Affitins are small, yet robust and easy to produce, proteins derived from the archaeal extremophilic "7kDa DNA-binding" protein family. By means of combinatorial protein engineering and ribosome display selection techniques, Affitins have shown to bind a diversity of targets. In this work, two previously developed Affitins (anti-lysozyme and anti-IgG) were immobilized onto magnetic particles to assess their potential for protein purification by magnetic fishing. The optimal lysozyme and human IgG binding conditions yielded 58mg lysozyme/g support and 165mgIgG/g support, respectively. The recovery of proteins was possible in high yield (≥95%) and with high purity, namely ≥95% and 81%, when recovering lysozyme from Escherichia coli supernatant and IgG from human plasma, respectively. Static binding studies indicated affinity constants of 5.0×10(4)M(-1) and 9.3×10(5)M(-1) for the anti-lysozyme and anti-IgG magnetic supports. This work demonstrated that Affitins, which can be virtually evolved for any protein of interest, can be coupled onto magnetic particles creating novel affinity adsorbents for purification by magnetic fishing. PMID:27342136

  8. Relative Binding Affinities of Monolignols to Horseradish Peroxidase.

    PubMed

    Sangha, Amandeep K; Petridis, Loukas; Cheng, Xiaolin; Smith, Jeremy C

    2016-08-11

    Monolignol binding to the peroxidase active site is the first step in lignin polymerization in plant cell walls. Using molecular dynamics, docking, and free energy perturbation calculations, we investigate the binding of monolignols to horseradish peroxidase C. Our results suggest that p-coumaryl alcohol has the strongest binding affinity followed by sinapyl and coniferyl alcohol. Stacking interactions between the monolignol aromatic rings and nearby phenylalanine residues play an important role in determining the calculated relative binding affinities. p-Coumaryl and coniferyl alcohols bind in a pose productive for reaction in which a direct H-bond is formed between the phenolic -OH group and a water molecule (W2) that may facilitate proton transfer during oxidation. In contrast, in the case of sinapyl alcohol there is no such direct interaction, the phenolic -OH group instead interacting with Pro139. Since proton and electron transfer is the rate-limiting step in monolignol oxidation by peroxidase, the binding pose (and thus the formation of near attack conformation) appears to play a more important role than the overall binding affinity in determining the oxidation rate. PMID:27447548

  9. Single-cell measurement of red blood cell oxygen affinity

    PubMed Central

    Di Caprio, Giuseppe; Stokes, Chris; Higgins, John M.; Schonbrun, Ethan

    2015-01-01

    Oxygen is transported throughout the body by hemoglobin (Hb) in red blood cells (RBCs). Although the oxygen affinity of blood is well-understood and routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of RBC volume and Hb concentration are taken millions of times per day by clinical hematology analyzers, and they are important factors in determining the health of the hematologic system. To better understand the variability and determinants of oxygen affinity on a cellular level, we have developed a system that quantifies the oxygen saturation, cell volume, and Hb concentration for individual RBCs in high throughput. We find that the variability in single-cell saturation peaks at an oxygen partial pressure of 2.9%, which corresponds to the maximum slope of the oxygen–Hb dissociation curve. In addition, single-cell oxygen affinity is positively correlated with Hb concentration but independent of osmolarity, which suggests variation in the Hb to 2,3-diphosphoglycerate (2–3 DPG) ratio on a cellular level. By quantifying the functional behavior of a cellular population, our system adds a dimension to blood cell analysis and other measurements of single-cell variability. PMID:26216973

  10. Pepsin-modified chiral monolithic column for affinity capillary electrochromatography.

    PubMed

    Hong, Tingting; Chi, Cuijie; Ji, Yibing

    2014-11-01

    Pepsin-modified affinity monolithic capillary electrochromatography, a novel microanalysis system, was developed by the covalent bonding of pepsin on silica monolith. The column was successfully applied in the chiral separation of (±)-nefopam. Furthermore, the electrochromatographic performance of the pepsin-functionalized monolith for enantiomeric analysis was evaluated in terms of protein content, pH of running buffer, sample volume, buffer concentration, applied voltage, and capillary temperature. The relative standard deviation (%RSD) values of retention time (intraday <0.53, n = 10; interday <0.53, n = 10; column-to-column <0.70, n = 20; and batch-to-batch <0.80, n = 20) indicated satisfactory stability of these columns. No appreciable change was observed in retention and resolution for chiral recognition of (±)-nefopam in 50 days with 100 injections. The proteolytic activity of this stationary phase was further characterized with bovine serum albumin as substrate for online protein digestion. As for monolithic immobilized enzyme reactor, successive protein injections confirmed both the operational stability and ability to reuse the bioreactor for at least 20 digestions. It implied that the affinity monolith used in this research opens a new path of exploring particularly versatile class of enzymes to develop enzyme-modified affinity capillary monolith for enantioseparation. PMID:25146884

  11. Comparison of Inlet Geometry in Microfluidic Cell Affinity Chromatography

    PubMed Central

    Li, Peng; Tian, Yu; Pappas, Dimitri

    2011-01-01

    Cell separation based on microfluidic affinity chromatography is a widely used methodology in cell analysis research when rapid separations with high purity are needed. Several successful examples have been reported with high separation efficiency and purity; however, cell capture at the inlet area and inlet design has not been extensively described or studied. The most common inlets—used to connect the microfluidic chip to pumps, tubing, etc—are vertical (top-loading) inlets and parallel (in-line) inlets. In this work, we investigated the cell capture behavior near the affinity chip inlet area and compared the different performance of vertical inlet devices and parallel inlet devices. Vertical inlet devices showed significant cell capture capability near the inlet area, which led to the formation of cell blockages as the separation progressed. Cell density near the inlet area was much higher than the remaining channel, while for parallel inlet chips cell density at the inlet area was similar to the rest of the channel. In this paper, we discuss the effects of inlet type on chip fabrication, nonspecific binding, cell capture efficiency, and separation purity. We also discuss the possibility of using vertical inlets in negative selection separations. Our findings show that inlet design is critical and must be considered when fabricating cell affinity microfluidic devices. PMID:21207967

  12. Negative Enrichment of Target Cells by Microfluidic Affinity Chromatography

    PubMed Central

    Li, Peng; Gao, Yan; Pappas, Dimitri

    2011-01-01

    A three-dimensional microfluidic channel was developed for high purity cell separations. This system featured high capture affinity using multiple vertical inlets to an affinity surface. In cell separations, positive selection (capture of the target cell) is usually employed. Negative enrichment, the capture of non-target cells and elution of target cells, has distinct advantages over positive selection. In negative enrichment, target cells are not labeled, and are not subjected to strenuous elution conditions or dilution. As a result, negative enrichment systems are amenable to multi-step processes in microfluidic systems. In previous work, we reported cell capture enhancement effects at vertical inlets to the affinity surface. In this study, we designed a chip that has multiple vertical and horizontal channels, forming a three-dimensional separation system. Enrichment of target cells showed separation purities of 92-96%, compared with straight-channel systems (77% purity). A parallelized chip was also developed for increased sample throughput. A two-channel showed similar separation purity with twice the sample flow rate. This microfluidic system, featuring high separation purity, ease of fabrication and use, is suitable for cell separations when subsequent analysis of target cells is required. PMID:21939198

  13. Supramolecular Affinity Chromatography for Methylation-Targeted Proteomics.

    PubMed

    Garnett, Graham A E; Starke, Melissa J; Shaurya, Alok; Li, Janessa; Hof, Fraser

    2016-04-01

    Proteome-wide studies of post-translationally methylated species using mass spectrometry are complicated by high sample diversity, competition for ionization among peptides, and mass redundancies. Antibody-based enrichment has powered methylation proteomics until now, but the reliability, pan-specificity, polyclonal nature, and stability of the available pan-specific antibodies are problematic and do not provide a standard, reliable platform for investigators. We have invented an anionic supramolecular host that can form host-guest complexes selectively with methyllysine-containing peptides and used it to create a methylysine-affinity column. The column resolves peptides on the basis of methylation-a feat impossible with a comparable commercial cation-exchange column. A proteolyzed nuclear extract was separated on the methyl-affinity column prior to standard proteomics analysis. This experiment demonstrates that such chemical methyl-affinity columns are capable of enriching and improving the analysis of methyllysine residues from complex protein mixtures. We discuss the importance of this advance in the context of biomolecule-driven enrichment methods. PMID:26973166

  14. Protein purification by aminosquarylium cyanine dye-affinity chromatography.

    PubMed

    Graça, Vânia C; Sousa, Fani; Santos, Paulo F; Almeida, Paulo S

    2015-01-01

    Affinity chromatography (AC) is one of the most important techniques for the separation and purification of biomolecules, being probably the most selective technique for protein purification. It is based on unique specific reversible interactions between the target molecule and a ligand. In this affinity interaction, the choice of the ligand is extremely important for the success of the purification protocol. The growing interest in AC has motivated an intense research effort toward the development of materials able to overcome the disadvantages of conventional natural ligands, namely their high cost and chemical and biological lability. In this context, synthetic dyes have emerged, in recent decades, as a promising alternative to biological ligands. Herein, detailed protocols for the assembling of a new chromatographic dye-ligand affinity support bearing an immobilized aminosquarylium cyanine dye on an agarose-based matrix (Sepharose CL-6B) and for the separation of a mixture o f three standard proteins: lysozyme, α-chymotrypsin, and trypsin are provided. PMID:25749942

  15. Affinity approaches in RNAi-based therapeutics purification.

    PubMed

    Pereira, Patrícia; Queiroz, João A; Figueiras, Ana; Sousa, Fani

    2016-05-15

    The recent investigation on RNA interference (RNAi) related mechanisms and applications led to an increased awareness of the importance of RNA in biology. Nowadays, RNAi-based technology has emerged as a potentially powerful tool for silencing gene expression, being exploited to develop new therapeutics for treating a vast number of human disease conditions, as it is expected that this technology can be translated onto clinical applications in a near future. This approach makes use of a large number of small (namely short interfering RNAs, microRNAs and PIWI-interacting RNAs) and long non-coding RNAs (ncRNAs), which are likely to have a crucial role as the next generation therapeutics. The commercial and biomedical interest in these RNAi-based therapy applications have fostered the need to develop innovative procedures to easily and efficiently purify RNA, aiming to obtain the final product with high purity degree, good quality and biological activity. Recently, affinity chromatography has been applied to ncRNAs purification, in view of the high specificity. Therefore, this article intends to review the biogenesis pathways of regulatory ncRNAs and also to discuss the most significant and recent developments as well as applications of affinity chromatography in the challenging task of purifying ncRNAs. In addition, the importance of affinity chromatography in ncRNAs purification is addressed and prospects for what is forthcoming are presented. PMID:26830537

  16. The Purification of a Blood Group A Glycoprotein: An Affinity Chromatography Experiment.

    ERIC Educational Resources Information Center

    Estelrich, J.; Pouplana, R.

    1988-01-01

    Describes a purification process through affinity chromatography necessary to obtain specific blood group glycoproteins from erythrocytic membranes. Discusses the preparation of erythrocytic membranes, extraction of glycoprotein from membranes, affinity chromatography purification, determination of glycoproteins, and results. (CW)

  17. Gas-phase lithium cation affinity of glycine.

    PubMed

    Bourcier, Sophie; Chiaa, Ru Xuan; Mimbong, Rosa Ngo Biboum; Bouchoux, Guy

    2015-01-01

    The gas-phase lithium cation binding thermochemistry of glycine has been determined theoretically by quantum chemical calculations at the G4 level and experimentally by the extended kinetic method using electrospray ionization quadrupole time-of-flight tandem mass spectrometry. The lithium cation affinity of glycine, ∆(Li)H°(298)(GLY), i.e. the∆(Li)H°(298) of the reaction GlyLi(+)→ Gly + Li(+)) given by the G4 method is equal to 241.4 kJ.mol(-1) if only the most stable conformer of glycine is considered or to 242.3 kJ.mol(-1) if the 298K equilibrium mixture of neutral conformers is included in the calculation. The ∆(Li)H°(298)(GLY) deduced from the extended kinetic method is obviously dependent on the choice of the Li(+) affinity scale, thus∆(Li)H°(298)(GLY) is equal to 228.7±0.9(2.0) kJ.mol(- 1) if anchored to the recently re-evaluated lithium cation affinity scale but shifted to 235.4±1.0 kJ.mol(-1) if G4 computed lithium cation affinities of the reference molecules is used. This difference of 6.3 kJ.mol(-1) may originate from a compression of the experimental lithium affinity scale in the high ∆(Li)H°(298) region. The entropy change associated with the reaction GlyLi(+)→Gly + Li(+) reveals a gain of approximately 15 J.mol(-) 1.K(-1) with respect to monodentate Li(+) acceptors. The origin of this excess entropy is attributed to the bidentate interaction between the Li(+) cation and both the carbonyl oxygen and the nitrogen atoms of glycine. The computed G4 Gibbs free energy,∆(Li)G°(298)(GLY) is equal to 205.3 kJ.mol(-1), a similar result, 201.0±3.4 kJ.mol(-1), is obtained from the experiment if the∆(Li)G°(298) of the reference molecules is anchored on the G4 results. PMID:26307695

  18. Robust adaptive control for a class of uncertain non-affine nonlinear systems using affine-type neural networks

    NASA Astrophysics Data System (ADS)

    Zhao, Shitie; Gao, Xianwen

    2016-08-01

    A robust adaptive control is proposed for a class of single-input single-output non-affine nonlinear systems. In order to approximate the unknown nonlinear function, a novel affine-type neural network is used, and then to compensate the approximation error and external disturbance a robust control term is employed. By Lyapunov stability analysis for the closed-loop system, it is proved that tracking errors asymptotically converge to zero. Moreover, an observer is designed to estimate the system states because all the states may not be available for measurements. Furthermore, the adaptation laws of neural networks and the robust controller are given out based on the Lyapunov stability theory. Finally, two simulation examples are presented to demonstrate the effectiveness of the proposed control method.

  19. A comparison of binding surfaces for SPR biosensing using an antibody-antigen system and affinity distribution analysis

    PubMed Central

    Zhao, Huaying; Gorshkova, Inna I.; Fu, Gregory L.; Schuck, Peter

    2013-01-01

    The application of optical biosensors in the study of macromolecular interactions requires immobilization of one binding partner to the surface. It is often highly desirable that the immobilization is uniform and does not affect the thermodynamic and kinetic binding parameters to soluble ligands. To achieve this goal, a variety of sensor surfaces, coupling strategies and surface chemistries are available. Previously, we have introduced a technique for increasing the level of detail on the immobilized sites beyond an average affinity by determining the distribution of affinities and kinetic rate constants from families of binding and dissociation traces acquired at different concentrations of soluble ligand. In the present work, we explore how this affinity distribution analysis can be useful in the assessment and optimization of surface immobilization. With this goal, using an antibody-antigen interaction as a model system, we study the activity, thermodynamic and kinetic binding parameters, and heterogeneity of surface sites produced with different commonly used sensor surfaces, at different total surface densities and with direct immobilization or affinity capture. PMID:23270815

  20. Evaluation and optimization of the metal-binding properties of a complex ligand for immobilized metal affinity chromatography.

    PubMed

    Chen, Bin; Li, Rong; Li, Shiyu; Chen, Xiaoli; Yang, Kaidi; Chen, Guoliang; Ma, Xiaoxun

    2016-02-01

    The simultaneous determination of two binding parameters for metal ions on an immobilized metal affinity chromatography column was performed by frontal chromatography. In this study, the binding parameters of Cu(2+) to l-glutamic acid were measured, the metal ion-binding characteristics of the complex ligand were evaluated. The linear correlation coefficients were all greater than 99%, and the relative standard deviations of two binding parameters were 0.58 and 0.059%, respectively. The experiments proved that the frontal chromatography method was accurate, reproducible, and could be used to determine the metal-binding parameters of the affinity column. The effects of buffer pH, type, and concentration on binding parameters were explored by uniform design experiment. Regression, matching and residual analyses of the models were performed. Meanwhile, the optimum-binding conditions of Cu(2+) on the l-glutamic acid-silica column were obtained. Under these binding conditions, observations and regression values of two parameters were similar, and the observation values were the best. The results demonstrated that high intensity metal affinity column could be effectively prepared by measuring and evaluating binding parameters using frontal chromatography combined with a uniform design experiment. The present work provided a new mode for evaluating and preparing immobilized metal affinity column with good metal-binding behaviors. PMID:26632098

  1. Binding of bisbenzylisoquinoline alkaloids to phosphatidylcholine vesicles and alveolar macrophages: relationship between binding affinity and antifibrogenic potential of these drugs.

    PubMed

    Ma, J K; Mo, C G; Malanga, C J; Ma, J Y; Castranova, V

    1991-01-01

    A group of bisbenzylisoquinoline alkaloids has been shown to exhibit various degrees of effectiveness in preventing silica-induced fibrosis in animal models. The objective of the present study was to characterize the binding of several of these alkaloids to phosphatidylcholine vesicles and rat alveolar macrophages using fluorometric and equilibrium dialysis methods, respectively. The lipid binding affinity of these alkaloids was found to depend upon several structural factors including hydrophobic substitutions, chiral configurations, and double oxygen bridge-restricted confirmation of the benzylisoquinoline moieties. Tetrandrine, which is a highly effective agent in preventing fibrosis, showed strong binding to both lipid vesicles and alveolar macrophages. In contrast, certain analogues of tetrandrine such as curine and tubocurine, which have little or no effect on silicosis, exhibited only weak binding to lipid vesicles and almost no binding to cells. The moderate binding affinity of fangchinoline to vesicles and cells corresponded to a moderate effectiveness of the compound as an antifibrogenic agent. Methoxyadiantifoline, an alkaloid of unknown antifibrogenic potential, also exhibited high binding affinities for lipid and cells. In conclusion, the results of these studies indicate that alveolar macrophages exhibit large binding capacities for certain members of this class of bisbenzylisoquinoline alkaloids. A positive correlation was observed between binding affinity to alveolar macrophages and the reported antifibrotic potency of these compounds. These data also suggest that the ability of these drugs to interact with alveolar macrophages may be a key step in inhibition of the progression of silica-induced pulmonary disease. PMID:1663032

  2. A versatile polypeptide platform for integrated recognition and reporting: affinity arrays for protein-ligand interaction analysis.

    PubMed

    Enander, Karin; Dolphin, Gunnar T; Liedberg, Bo; Lundström, Ingemar; Baltzer, Lars

    2004-05-17

    A molecular platform for protein detection and quantification is reported in which recognition has been integrated with direct monitoring of target-protein binding. The platform is based on a versatile 42-residue helix-loop-helix polypeptide that dimerizes to form four-helix bundles and allows site-selective modification with recognition and reporter elements on the side chains of individually addressable lysine residues. The well-characterized interaction between the model target-protein carbonic anhydrase and its inhibitor benzenesulfonamide was used for a proof-of-concept demonstration. An affinity array was designed where benzenesulfonamide derivatives with aliphatic or oligoglycine spacers and a fluorescent dansyl reporter group were introduced into the scaffold. The affinities of the array members for human carbonic anhydrase II (HCAII) were determined by titration with the target protein and were found to be highly affected by the properties of the spacers (dissociation constant Kd=0.02-3 microM). The affinity of HCAII for acetazolamide (Kd=4 nM) was determined in a competition experiment with one of the benzenesulfonamide array members to address the possibility of screening substance libraries for new target-protein binders. Also, successful affinity discrimination between different carbonic anhydrase isozymes highlighted the possibility of performing future isoform-expression profiling. Our platform is predicted to become a flexible tool for a variety of biosensor and protein-microarray applications within biochemistry, diagnostics and pharmaceutical chemistry. PMID:15146511

  3. Thiochrome enhances acetylcholine affinity at muscarinic M4 receptors: receptor subtype selectivity via cooperativity rather than affinity.

    PubMed

    Lazareno, S; Dolezal, V; Popham, A; Birdsall, N J M

    2004-01-01

    Thiochrome (2,7-dimethyl-5H-thiachromine-8-ethanol), an oxidation product and metabolite of thiamine, has little effect on the equilibrium binding of l-[3H]N-methyl scopolamine ([3H]NMS) to the five human muscarinic receptor subtypes (M1-M5) at concentrations up to 0.3 mM. In contrast, it inhibits [3H]NMS dissociation from M1 to M4 receptors at submillimolar concentrations and from M5 receptors at 1 mM. These results suggest that thiochrome binds allosterically to muscarinic receptors and has approximately neutral cooperativity with [3H]NMS at M1 to M4 and possibly M5 receptors. Thiochrome increases the affinity of acetylcholine (ACh) 3- to 5-fold for inhibiting [3H]NMS binding to M4 receptors but has no effect on ACh affinity at M1 to M3 or M5 receptors. Thiochrome (0.1 mM) also increases the direct binding of [3H]ACh to M4 receptors but decreases it slightly at M2 receptors. In agreement with the binding data, thiochrome does not affect the potency of ACh for stimulating the binding of guanosine 5'-O-(3-[35S]thiotriphosphate) ([35S]GTPgammaS) to membranes containing M1 to M3 receptors, but it increases ACh potency 3.5-fold at M4 receptors. It also selectively reduces the release of [3H]ACh from potassium-stimulated slices of rat striatum, which contain autoinhibitory presynaptic M4 receptors, but not from hippocampal slices, which contain presynaptic M2 receptors. We conclude that thiochrome is a selective M4 muscarinic receptor enhancer of ACh affinity and has neutral cooperativity with ACh at M1 to M3 receptors; it therefore demonstrates a powerful new form of selectivity, "absolute subtype selectivity", which is derived from cooperativity rather than from affinity. PMID:14722259

  4. Building the bacterial orisome: high affinity DnaA recognition plays a role in setting the conformation of oriC DNA

    PubMed Central

    Kaur, Gulpreet; Vora, Mansi P.; Czerwonka, Christopher A.; Rozgaja, Tania A.; Grimwade, Julia E.; Leonard, Alan C.

    2014-01-01

    Summary During assembly of the E. coli pre-replicative complex (pre-RC), initiator DnaA oligomers are nucleated from three widely separated high affinity DnaA recognition sites in oriC. Oligomer assembly is then guided by low affinity DnaA recognition sites, but is also regulated by a switch-like conformational change in oriC mediated by sequential binding of two DNA bending proteins, Fis and IHF, serving as inhibitor and activator, respectively. Although their recognition sites are separated by up to 90 bp, Fis represses IHF binding and weak DnaA interactions until accumulating DnaA displaces Fis from oriC. It remains unclear whether high affinity DnaA binding plays any role in Fis repression at a distance and it is also not known whether all high affinity DnaA recognition sites play an equivalent role in oligomer formation. To examine these issues, we developed origin-selective recombineering methods to mutate E. coli chromosomal oriC. We found that, although oligomers were assembled in the absence of any individual high affinity DnaA binding site, loss of DnaA binding at peripheral sites eliminated Fis repression, and made binding of both Fis and IHF essential. We propose a model in which interaction of DnaA molecules at high affinity sites regulates oriC DNA conformation. PMID:24443848

  5. The neonatal Fc receptor (FcRn) binds independently to both sites of the IgG homodimer with identical affinity

    PubMed Central

    Abdiche, Yasmina Noubia; Yeung, Yik Andy; Chaparro-Riggers, Javier; Barman, Ishita; Strop, Pavel; Chin, Sherman Michael; Pham, Amber; Bolton, Gary; McDonough, Dan; Lindquist, Kevin; Pons, Jaume; Rajpal, Arvind

    2015-01-01

    The neonatal Fc receptor (FcRn) is expressed by cells of epithelial, endothelial and myeloid lineages and performs multiple roles in adaptive immunity. Characterizing the FcRn/IgG interaction is fundamental to designing therapeutic antibodies because IgGs with moderately increased binding affinities for FcRn exhibit superior serum half-lives and efficacy. It has been hypothesized that 2 FcRn molecules bind an IgG homodimer with disparate affinities, yet their affinity constants are inconsistent across the literature. Using surface plasmon resonance biosensor assays that eliminated confounding experimental artifacts, we present data supporting an alternate hypothesis: 2 FcRn molecules saturate an IgG homodimer with identical affinities at independent sites, consistent with the symmetrical arrangement of the FcRn/Fc complex observed in the crystal structure published by Burmeister et al. in 1994. We find that human FcRn binds human IgG1 with an equilibrium dissociation constant (KD) of 760 ± 60 nM (N = 14) at 25°C and pH 5.8, and shows less than 25% variation across the other human subtypes. Human IgG1 binds cynomolgus monkey FcRn with a 2-fold higher affinity than human FcRn, and binds both mouse and rat FcRn with a 10-fold higher affinity than human FcRn. FcRn/IgG interactions from multiple species show less than a 2-fold weaker affinity at 37°C than at 25°C and appear independent of an IgG's variable region. Our in vivo data in mouse and rat models demonstrate that both affinity and avidity influence an IgG's serum half-life, which should be considered when choosing animals, especially transgenic systems, as surrogates. PMID:25658443

  6. Shark Attack: high affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation.

    PubMed

    Zielonka, Stefan; Weber, Niklas; Becker, Stefan; Doerner, Achim; Christmann, Andreas; Christmann, Christine; Uth, Christina; Fritz, Janine; Schäfer, Elena; Steinmann, Björn; Empting, Martin; Ockelmann, Pia; Lierz, Michael; Kolmar, Harald

    2014-12-10

    A novel method for stepwise in vitro affinity maturation of antigen-specific shark vNAR domains is described that exclusively relies on semi-synthetic repertoires derived from non-immunized sharks. Target-specific molecules were selected from a CDR3-randomized bamboo shark (Chiloscyllium plagiosum) vNAR library using yeast surface display as platform technology. Various antigen-binding vNAR domains were easily isolated by screening against several therapeutically relevant antigens, including the epithelial cell adhesion molecule (EpCAM), the Ephrin type-A receptor 2 (EphA2), and the human serine protease HTRA1. Affinity maturation was demonstrated for EpCAM and HTRA1 by diversifying CDR1 of target-enriched populations which allowed for the rapid selection of nanomolar binders. EpCAM-specific vNAR molecules were produced as soluble proteins and more extensively characterized via thermal shift assays and biolayer interferometry. Essentially, we demonstrate that high-affinity binders can be generated in vitro without largely compromising the desirable high thermostability of the vNAR scaffold. PMID:24862193

  7. Biological affinities and adaptations of Bronze Age Bactrians: IV. A craniometric investigation of Bactrian origins.

    PubMed

    Hemphill, B E

    1999-02-01

    Discovery of a previously unknown Bronze Age civilization (Oxus civilization) centered on the oases of Central Asia revealed the presence of large, preplanned urban centers immediately above sterile soil. Given the absence of local antecedents, the sudden appearance and proliferation of these Oxus civilization urban centers in the oases of Bactria and Margiana immediately raised the issue of where the inhabitants of these urban centers came from. Three hypotheses have been offered by archaeologists to account for the origins of Oxus civilization populations. These include the early influence model, the late colonization model, and the trichotomy model. Eleven craniometric variables from 12 Aeneolithic and Bronze Age samples, encompassing 657 adults from Central Asia, Iran, and the Indus Valley, are compared to test which if any of these hypotheses are supported by the pattern of phenetic affinities possessed by the Oxus civilization inhabitants of the north Bactrian oasis. Craniometric differences between samples are compared with Mahalanobis generalized distance (d2), and patterns of phenetic affinity are assessed with two types of cluster analysis (WPGMA, neighbor-joining method), multidimensional scaling, and principal coordinates analysis. Results of this analysis provide no support for either the late colonization model or the trichotomy model but do offer some support for the early influence model. Nevertheless, it is clear that the early influence model fails to account for a shift in interregional contacts, perhaps from western China to the north around 2000 BC, that appears to have played a major role in the origins of the Oxus civilization inhabitants of the north Bactrian oasis. PMID:9988380

  8. PHARMACEUTICAL AND BIOMEDICAL APPLICATIONS OF AFFINITY CHROMATOGRAPHY: RECENT TRENDS AND DEVELOPMENTS

    PubMed Central

    Hage, David S.; Anguizola, Jeanethe A.; Bi, Cong; Li, Rong; Matsuda, Ryan; Papastavros, Efthimia; Pfaunmiller, Erika; Vargas, John; Zheng, Xiwei

    2012-01-01

    Affinity chromatography is a separation technique that has become increasingly important in work with biological samples and pharmaceutical agents. This method is based on the use of a biologically-related agent as a stationary phase to selectively retain analytes or to study biological interactions. This review discusses the basic principles behind affinity chromatography and examines recent developments that have occurred in the use of this method for biomedical and pharmaceutical analysis. Techniques based on traditional affinity supports are discussed, but an emphasis is placed on methods in which affinity columns are used as part of HPLC systems or in combination with other analytical methods. General formats for affinity chromatography that are considered include step elution schemes, weak affinity chromatography, affinity extraction and affinity depletion. Specific separation techniques that are examined include lectin affinity chromatography, boronate affinity chromatography, immunoaffinity chromatography, and immobilized metal ion affinity chromatography. Approaches for the study of biological interactions by affinity chromatography are also presented, such as the measurement of equilibrium constants, rate constants, or competition and displacement effects. In addition, related developments in the use of immobilized enzyme reactors, molecularly imprinted polymers, dye ligands and aptamers are briefly considered. PMID:22305083

  9. Thermoviscoplastic behaviors of anisotropic shape memory elastomeric composites for cold programmed non-affine shape change

    NASA Astrophysics Data System (ADS)

    Mao, Yiqi; Robertson, Jaimee M.; Mu, Xiaoming; Mather, Patrick T.; Jerry Qi, H.

    2015-12-01

    Shape memory polymers (SMPs) can fix a temporary shape and recover their permanent shape upon activation by an external stimulus. Most SMPs require programming at above their transition temperatures, normally well above the room temperature. In addition, most SMPs are programmed into shapes that are affine to the high temperature deformation. Recently, a cold-programmed anisotropic shape memory elastomeric composite was developed and showed interesting low temperature stretching induced shape memory behavior. There, simple, uniaxial stretching at low temperature transformed the composites into curled temporary shapes upon unloading. The exact geometry of the curled state depended on the microstructure of the composite, and the curled shape showed no affinity to the deformed shape. Heating the sample recovered the sample back to its original shape. This new composite consisted of an elastomeric matrix reinforced by aligned amorphous polymer fibers. By utilizing the plastic-like behavior of the amorphous polymer phase at low temperatures, a temporary shape could be fixed upon unloading since the induced plastic-like strain resists the recovery of the elastomer matrix. After heating to a high temperature, the permanent shape was recovered when the amorphous polymer softened and the elastomer matrix contracted. To set a theoretical foundation for capturing the cold-programmed shape memory effects and the dramatic non-affine shape change of this composite, a 3D anisotropic thermoviscoelastic constitutive model is developed in this paper. In this model, the matrix is modeled as a hyperelastic solid, and the amorphous phase of the fibrous mat is considered as a nonlinear thermoviscoplastic solid, whose viscous flow resistance is sensitive to both temperature and stress. The plastic-deformation like behavior demonstrated in the fiber is treated as nonlinear viscoplasticity with extremely high viscosity or long relaxation time at zero-stress state at low temperature. The

  10. Domain Selection in Metallothionein 1A: Affinity-Controlled Mechanisms of Zinc Binding and Cadmium Exchange.

    PubMed

    Pinter, Tyler B J; Irvine, Gordon W; Stillman, Martin J

    2015-08-18

    Mammalian metallothioneins (MTs) are small, metal binding proteins implicated in cellular metal ion homeostasis and heavy metal detoxification. Divalent, metal-saturated MTs form two distinct domains; the N-terminal β domain binds three metals using nine Cys residues, and the C-terminal α domain binds four metals with 11 Cys residues. Domain selection during zinc binding and cadmium exchange to human MT1A was examined using a series of competition reactions with mixtures of the isolated domain fragments. These experiments were conducted at two biologically significant pH conditions where MTs exist in vivo. Neither zinc binding nor cadmium exchange showed any significant degree of specificity or selectivity based on detailed analysis of electrospray ionization mass spectrometric and circular dichroic data. Under acidic conditions, zinc binding and cadmium exchange showed slight α domain selectivity because of the increased preference for cooperative clustering of the α domain. Modeling of the reactions showed that at both physiological (7.4) and acidic (5.8) pHs, zinc binding and cadmium exchanges occur essentially randomly between the two fragments. The metal binding affinity distributions between the domain fragments are comingled and not significantly separated as required for a domain specific mechanism. The models show rather that the order of the binding events follows the order of the binding affinities that are distributed across both domains and that this can be considered quantitatively by the KF(Cd)/KF(Zn) binding constant ratio for each metal bound. PMID:26167879

  11. Metal-Mediated Affinity and Orientation Specificity in a Computationally Designed Protein Homodimer

    SciTech Connect

    Der, Bryan S.; Machius, Mischa; Miley, Michael J.; Mills, Jeffrey L.; Szyperski, Thomas; Kuhlman, Brian

    2015-10-15

    Computationally designing protein-protein interactions with high affinity and desired orientation is a challenging task. Incorporating metal-binding sites at the target interface may be one approach for increasing affinity and specifying the binding mode, thereby improving robustness of designed interactions for use as tools in basic research as well as in applications from biotechnology to medicine. Here we describe a Rosetta-based approach for the rational design of a protein monomer to form a zinc-mediated, symmetric homodimer. Our metal interface design, named MID1 (NESG target ID OR37), forms a tight dimer in the presence of zinc (MID1-zinc) with a dissociation constant <30 nM. Without zinc the dissociation constant is 4 {micro}M. The crystal structure of MID1-zinc shows good overall agreement with the computational model, but only three out of four designed histidines coordinate zinc. However, a histidine-to-glutamate point mutation resulted in four-coordination of zinc, and the resulting metal binding site and dimer orientation closely matches the computational model (C{alpha} rmsd = 1.4 {angstrom}).

  12. ANALYSIS OF DRUG-PROTEIN BINDING BY ULTRAFAST AFFINITY CHROMATOGRAPHY USING IMMOBILIZED HUMAN SERUM ALBUMIN

    PubMed Central

    Mallik, Rangan; Yoo, Michelle J.; Briscoe, Chad J.; Hage, David S.

    2010-01-01

    Human serum albumin (HSA) was explored for use as a stationary phase and ligand in affinity microcolumns for the ultrafast extraction of free drug fractions and the use of this information for the analysis of drug-protein binding. Warfarin, imipramine, and ibuprofen were used as model analytes in this study. It was found that greater than 95% extraction of all these drugs could be achieved in as little as 250 ms on HSA microcolumns. The retained drug fraction was then eluted from the same column under isocratic conditions, giving elution in less than 40 s when working at 4.5 mL/min. The chromatographic behavior of this system gave a good fit with that predicted by computer simulations based on a reversible, saturable model for the binding of an injected drug with immobilized HSA. The free fractions measured by this method were found to be comparable to those determined by ultrafiltration, and equilibrium constants estimated by this approach gave good agreement with literature values. Advantages of this method include its speed and the relatively low cost of microcolumns that contain HSA. The ability of HSA to bind many types of drugs also creates the possibility of using the same affinity microcolumn to study and measure the free fractions for a variety of pharmaceutical agents. These properties make this technique appealing for use in drug binding studies and in the high-throughput screening of new drug candidates. PMID:20227701

  13. DETECTION OF HETEROGENEOUS DRUG-PROTEIN BINDING BY FRONTAL ANALYSIS AND HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Tong, Zenghan; Joseph, K.S.; Hage, David S.

    2011-01-01

    This study examined the use of frontal analysis and high-performance affinity chromatography for detecting heterogeneous binding in biomolecular interactions, using the binding of acetohexamide with human serum albumin (HSA) as a model. It was found through the use of this model system and chromatographic theory that double-reciprocal plots could be used more easily than traditional isotherms for the initial detection of binding site heterogeneity. The deviations from linearity that were seen in double-reciprocal plots as a result of heterogeneity were a function of the analyte concentration, the relative affinities of the binding sites in the system and the amount of each type of site that was present. The size of these deviations was determined and compared under various conditions. Plots were also generated to show what experimental conditions would be needed to observe these deviations for general heterogeneous systems or for cases in which some preliminary information was available on the extent of binding heterogeneity. The methods developed in this work for the detection of binding heterogeneity are not limited to drug interactions with HSA but could be applied to other types of drug-protein binding or to additional biological systems with heterogeneous binding. PMID:21612784

  14. Correlation and prediction of adsorption capacity and affinity of aromatic compounds on carbon nanotubes.

    PubMed

    Wu, Wenhao; Yang, Kun; Chen, Wei; Wang, Wendi; Zhang, Jie; Lin, Daohui; Xing, Baoshan

    2016-01-01

    Adsorption of 22 nonpolar and polar aromatic compounds on 10 carbon nanotubes (CNTs) with various diameters, lengths and surface oxygen-containing group contents was investigated to develop predictive correlations for adsorption, using the isotherm fitting of Polanyi theory-based Dubinin-Ashtakhov (DA) model. Adsorption capacity of aromatic compounds on CNTs is negatively correlated with melting points of aromatic compounds, and surface oxygen-containing group contents and surface area ratios of mesopores to total pores of CNTs, but positively correlated with total surface area of CNTs. Adsorption affinity is positively correlated with solvatochromic parameters of aromatic compounds, independent of tube lengths and surface oxygen-containing group contents of CNTs, but negatively correlated with surface area ratios of mesopores to total pores of CNTs. The correlations of adsorption capacity and adsorption affinity with properties of both aromatic compounds and CNTs clearly have physical significance, can be used successfully with DA model to predict adsorption of aromatic compounds on CNTs from the well-known physiochemical properties of aromatic compounds (i.e., solvatochromic parameters, melting points) and CNTs (i.e., surface area and total acidic group contents), and thus can facilitate the environmental application of CNTs as sorbents and environmental risk assessment of both aromatic contaminants and CNTs. PMID:26521219

  15. Lipid-based nanoparticles with high binding affinity for amyloid-beta1-42 peptide.

    PubMed

    Gobbi, Marco; Re, Francesca; Canovi, Mara; Beeg, Marten; Gregori, Maria; Sesana, Silvia; Sonnino, Sandro; Brogioli, Doriano; Musicanti, Claudia; Gasco, Paolo; Salmona, Mario; Masserini, Massimo E

    2010-09-01

    The neurotoxic beta-amyloid peptide (Abeta), formed in anomalous amounts in Alzheimer's disease (AD), is released as monomer and then undergoes aggregation forming oligomers, fibrils and plaques in diseased brains. Abeta aggregates are considered as possible targets for therapy and/or diagnosis of AD. Since nanoparticles (NPs) are promising vehicles for imaging probes and therapeutic agents, we realized and characterized two types of NPs (liposomes and solid lipid nanoparticles, 145 and 76 nm average size, respectively) functionalized to target Abeta(1-42) with high affinity. Preliminary immunostaining studies identified anionic phospholipids [phosphatidic acid (PA) and cardiolipin (CL)] as suitable Abeta(1-42) ligands. PA/CL-functionalized, but not plain, NPs interacted with Abeta(1-42) aggregates as indicated by ultracentrifugation experiments, in which binding reaction occurred in solution, and by Surface Plasmon Resonance (SPR) experiments, in which NPs flowed onto immobilized Abeta(1-42). All these experiments were carried out in buffered saline. SPR studies indicated that, when exposed on NPs surface, PA/CL display very high affinity for Abeta(1-42) fibrils (22-60 nm), likely because of the occurrence of multivalent interactions which markedly decrease the dissociation of PA/CL NPs from Abeta. Noteworthy, PA/CL NPs did not bind to bovine serum albumin. The PA/CL NPs described in this work are endowed with the highest affinity for Abeta so far reported. These characteristics make our NPs a very promising vector for the targeted delivery of potential new diagnostic and therapeutic molecules to be tested in appropriate animal models. PMID:20553982

  16. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses

    PubMed Central

    Shorter, Shayla K.; Schnell, Frederick J.; McMaster, Sean R.; Pinelli, David F.; Andargachew, Rakieb; Evavold, Brian D.

    2016-01-01

    T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC) or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL), have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV) escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4) are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant. PMID:26915099

  17. Nickel accumulation in leaves, floral organs and rewards varies by serpentine soil affinity

    PubMed Central

    Meindl, George A.; Bain, Daniel J.; Ashman, Tia-Lynn

    2014-01-01

    Serpentine soils are edaphically stressful environments that host many endemic plant species. In particular, serpentine soils are high in several heavy metals (e.g. nickel, cobalt and chromium) and these high heavy metal concentrations are thought, in part, to lead to varying levels of plant adaptation and soil affinities (i.e. endemic vs. non-endemic plant species). It is unclear, however, whether serpentine endemics vs. non-endemics differ with respect to heavy metal uptake into either vegetative or reproductive organs. Here, we use nickel as a model to determine whether plant heavy metal uptake varies with the level of endemism in several non-hyperaccumulating species. Under controlled greenhouse conditions, we grew seven plant species from the Brassicaceae family that vary in their degrees of affinity to serpentine soil from low (indifferent) to medium (indicator) and high (endemic) in soil that was nickel supplemented or not. We quantified nickel concentrations in leaves, pistils, anthers, pollen and nectar. While nickel concentrations did not vary across organs or affinities when grown in control soils, under conditions of nickel supplementation endemic species had the lowest tissue concentrations of nickel, particularly when considering leaves and pistils, compared with indifferent/indicator species. Species indifferent to serpentines incorporated higher concentrations of nickel into reproductive organs relative to leaves, but this was not the case for indicator species and endemics where nickel concentration was similar in these organs. Our findings suggest that endemic species possess the ability to limit nickel uptake into above-ground tissues, particularly in reproductive organs where it may interfere with survival and reproduction. Indifferent species accumulated significantly more nickel into reproductive organs compared with leaves, which may limit their reproductive potential relative to endemic species when growing on serpentine soils. Additional

  18. AB-Bind: Antibody binding mutational database for computational affinity predictions.

    PubMed

    Sirin, Sarah; Apgar, James R; Bennett, Eric M; Keating, Amy E

    2016-02-01

    Antibodies (Abs) are a crucial component of the immune system and are often used as diagnostic and therapeutic agents. The need for high-affinity and high-specificity antibodies in research and medicine is driving the development of computational tools for accelerating antibody design and discovery. We report a diverse set of antibody binding data with accompanying structures that can be used to evaluate methods for modeling antibody interactions. Our Antibody-Bind (AB-Bind) database includes 1101 mutants with experimentally determined changes in binding free energies (ΔΔG) across 32 complexes. Using the AB-Bind data set, we evaluated the performance of protein scoring potentials in their ability to predict changes in binding free energies upon mutagenesis. Numerical correlations between computed and observed ΔΔG values were low (r = 0.16-0.45), but the potentials exhibited predictive power for classifying variants as improved vs weakened binders. Performance was evaluated using the area under the curve (AUC) for receiver operator characteristic (ROC) curves; the highest AUC values for 527 mutants with |ΔΔG| > 1.0 kcal/mol were 0.81, 0.87, and 0.88 using STATIUM, FoldX, and Discovery Studio scoring potentials, respectively. Some methods could also enrich for variants with improved binding affinity; FoldX and Discovery Studio were able to correctly rank 42% and 30%, respectively, of the 80 most improved binders (those with ΔΔG < -1.0 kcal/mol) in the top 5% of the database. This modest predictive performance has value but demonstrates the continuing need to develop and improve protein energy functions for affinity prediction. PMID:26473627

  19. New horizons in mouse immunoinformatics: reliable in silico prediction of mouse class I histocompatibility major complex peptide binding affinity.

    PubMed

    Hattotuwagama, Channa K; Guan, Pingping; Doytchinova, Irini A; Flower, Darren R

    2004-11-21

    Quantitative structure-activity relationship (QSAR) analysis is a main cornerstone of modern informatic disciplines. Predictive computational models, based on QSAR technology, of peptide-major histocompatibility complex (MHC) binding affinity have now become a vital component of modern day computational immunovaccinology. Historically, such approaches have been built around semi-qualitative, classification methods, but these are now giving way to quantitative regression methods. The additive method, an established immunoinformatics technique for the quantitative prediction of peptide-protein affinity, was used here to identify the sequence dependence of peptide binding specificity for three mouse class I MHC alleles: H2-D(b), H2-K(b) and H2-K(k). As we show, in terms of reliability the resulting models represent a significant advance on existing methods. They can be used for the accurate prediction of T-cell epitopes and are freely available online ( http://www.jenner.ac.uk/MHCPred). PMID:15534705

  20. Cloning and functional characterization of an Arabidopsis nitrate transporter gene that encodes a constitutive component of low-affinity uptake.

    PubMed Central

    Huang, N C; Liu, K H; Lo, H J; Tsay, Y F

    1999-01-01

    The Arabidopsis CHL1 (AtNRT1) gene encodes an inducible component of low-affinity nitrate uptake, which necessitates a "two-component" model to account for the constitutive low-affinity uptake observed in physiological studies. Here, we report the cloning and characterization of a CHL1 homolog, AtNRT1:2 (originally named NTL1), with data to indicate that this gene encodes a constitutive component of low-affinity nitrate uptake. Transgenic plants expressing antisense AtNRT1:2 exhibited reduced nitrate-induced membrane depolarization and nitrate uptake activities in assays with 10 mM nitrate. Furthermore, transgenic plants expressing antisense AtNRT1:2 in the chl1-5 background exhibited an enhanced resistance to chlorate (7 mM as opposed to 2 mM for the chl1-5 mutant). Kinetic analysis of AtNRT1:2-injected Xenopus oocytes yielded a K(m) for nitrate of approximately 5.9 mM. In contrast to CHL1, AtNRT1:2 was constitutively expressed before and after nitrate exposure (it was repressed transiently only when the level of CHL1 mRNA started to increase significantly), and its mRNA was found primarily in root hairs and the epidermis in both young (root tips) and mature regions of roots. We conclude that low-affinity systems of nitrate uptake, like high-affinity systems, are composed of inducible and constitutive components and that with their distinct functions, they are part of an elaborate nitrate uptake network in Arabidopsis. PMID:10449574