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Sample records for affine toda field

  1. Complex Affine Toda Theories and Soliton Solutions

    NASA Astrophysics Data System (ADS)

    Zhu, Zhiqing

    1995-01-01

    Toda field theories (TFT's) constitute a large class of integrable (1 + 1)-dimensional field theories that are relativistically invariant: included are conformal field theories and integrable deformations away from conformality. Because they are soluble, for example, by the inverse scattering method, and because they are related to many other areas of field theory, they have been studied extensively in recent years. Hirota's method is a straightforward procedure to obtain soliton solutions to non-linear integrable equations. In Hirota's method, one first writes the nonlinear equations in Hirota's bilinear form, and then expands the so called tau-functions as a power series in an arbitrary parameter. The power series terminates at some finite order, thus the solutions obtained are exact. For an N-soliton solution, the number of terms in the expansion grows exponentially with N, making direct calculation of N-soliton solutions difficult. We extend Hirota's one -parameter expansion to an N-parameter expansion. In the new expansion series, many terms are identical to those in the (N - 1)-soliton solutions, and new terms grow only linearly with N. Furthermore, we note that the expansion must terminate at some finite order, thus the vanishing of higher order terms can be used as constraints on these new terms. It turns out that these constraints can be used to determine the new terms completely. We used this extended Hirota's method to find N-soliton solutions for complex affine TFT's based on a simply-laced Kac-Moody algebra. Soliton solutions for non-simply-laced complex ATFT's can be obtained for those of simply-laced complex ATFT's by folding or twisting. Even though some soliton solutions have already been obtained for complex ATFT's by various methods, the physical implications of these solutions have not yet been thoroughly discussed. There are infinitely many distinct topological solitons in any given complex affine Toda field theory and most of them have complex

  2. 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.

  3. 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.

  4. Conformal field theory on affine Lie groups

    SciTech Connect

    Clubok, K.S.

    1996-04-01

    Working directly on affine Lie groups, we construct several new formulations of the WZW model, the gauged WZW model, and the generic affine-Virasoro action. In one formulation each of these conformal field theories (CFTs) is expressed as a one-dimensional mechanical system whose variables are coordinates on the affine Lie group. When written in terms of the affine group element, this formulation exhibits a two-dimensional WZW term. In another formulation each CFT is written as a two-dimensional field theory, with a three- dimensional WZW term, whose fields are coordinates on the affine group. On the basis of these equivalent formulations, we develop a translation dictionary in which the new formulations on the affine Lie group are understood as mode formulations of the conventional formulations on the Lie group. Using this dictionary, we also express each CFT as a three-dimensional field theory on the Lie group with a four-dimensional WZW term. 36 refs.

  5. 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.

  6. Vertex operator representation of the soliton tau functions in the An(1) Toda models by dressing transformations

    NASA Astrophysics Data System (ADS)

    Belich, H.; Cuba, G.; Paunov, R.

    1998-10-01

    We study the relation between the group-algebraic approach and the dressing symmetry one to the soliton solutions of the An(1) Toda field theory in 1+1 dimensions. Originally, solitons in the affine Toda models were found by Olive, Turok, and Underwood. Single solitons are created by exponentials of elements which ad-diagonalize the principal Heisenberg subalgebra. Alternatively, Babelon and Bernard exploited the dressing symmetry to reproduce the known expressions for the fundamental tau functions in the sine-Gordon model. In this paper we show the equivalence between these two methods to construct solitons in the An(1) Toda models.

  7. On Some Periodic Toda Lattices

    PubMed Central

    Kac, M.; Van Moerbeke, Pierre

    1975-01-01

    A discrete version of Floquet's theory is developed and applied to a system of non-linear differential equations related to the periodic Toda lattice. A special solution previously found by Toda is thus seen to fit into the formalism of inverse scattering problems. PMID:16592244

  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. Toda 3-point functions from topological strings II

    NASA Astrophysics Data System (ADS)

    Isachenkov, Mikhail; Mitev, Vladimir; Pomoni, Elli

    2016-08-01

    In [1] we proposed a formula for the 3-point structure constants of generic primary fields in the Toda field theory, derived using topological strings and the AGT-W correspondence from the partition functions of the non-Lagrangian T N theories on S 4. In this article, we obtain from it the well-known formula by Fateev and Litvinov and show that the degeneration on a first level of one of the three primary fields on the Toda side corresponds to a particular Higgsing of the T N theories.

  10. Field-Emission from Chemically Functionalized Diamond Surfaces: Does Electron Affinity Picture Work?

    NASA Astrophysics Data System (ADS)

    Miyamoto, Yoshiyuki; Miyazaki, Takehide; Takeuchi, Daisuke; Okushi, Hideyo; Yamasaki, Satoshi

    2014-03-01

    By means of the time-dependent density functional electron dynamics, we have revisited the field-emission efficiency of chemically functionalized diamond (100) surfaces. In order to achieve high efficiency and high (chemical) stability, proper chemical species are needed to terminate diamond surfaces. Hydrogen (H) termination is well known to achieve the negative electron affinity (NEA) of diamond surface which indeed enhances field emission performance than that of clean surface with positive electron affinity (PEA). Yet, the durability of H-terminated diamond surface was concerned for long-time operation of the field-emission. Meantime, oxidation, or hydroxyl (OH) termination was considered to achieve chemical stability of the surface but presence of oxygen (O) atom should reduce the emission efficiency. Recently, H- OH-co-terminated surface is reported as NEA and was expected to achieve both emission efficiency and chemical stability. However, our simulation showed that emission efficiency of the H- OH- co-terminated surface is much lower than clean surface with PEA, thus we note that the electron affinity cannot be a unique measure to determine the emission efficiency. In this talk, we introduce necessity of new concept to understand the emission efficiency which needs to know detailed potential profile from bulk to vacuum through surface, which is strongly dependent on the surface chemical functionalization. This work was supported by ALCA project conducted by Japan Science and Technology Agency.

  11. Electric Field Cancellation on Quartz by Rb Adsorbate-Induced Negative Electron Affinity

    NASA Astrophysics Data System (ADS)

    Sedlacek, J. A.; Kim, E.; Rittenhouse, S. T.; Weck, P. F.; Sadeghpour, H. R.; Shaffer, J. P.

    2016-04-01

    We investigate the (0001) surface of single crystal quartz with a submonolayer of Rb adsorbates. Using Rydberg atom electromagnetically induced transparency, we investigate the electric fields resulting from Rb adsorbed on the quartz surface, and measure the activation energy of the Rb adsorbates. We show that the adsorbed Rb induces negative electron affinity (NEA) on the quartz surface. The NEA surface allows low energy electrons to bind to the surface and cancel the electric field from the Rb adsorbates. Our results will be important for integrating Rydberg atoms into hybrid quantum systems, as fundamental probes of atom-surface interactions, and for studies of 2D electron gases bound to surfaces.

  12. Electric Field Cancellation on Quartz by Rb Adsorbate-Induced Negative Electron Affinity.

    PubMed

    Sedlacek, J A; Kim, E; Rittenhouse, S T; Weck, P F; Sadeghpour, H R; Shaffer, J P

    2016-04-01

    We investigate the (0001) surface of single crystal quartz with a submonolayer of Rb adsorbates. Using Rydberg atom electromagnetically induced transparency, we investigate the electric fields resulting from Rb adsorbed on the quartz surface, and measure the activation energy of the Rb adsorbates. We show that the adsorbed Rb induces negative electron affinity (NEA) on the quartz surface. The NEA surface allows low energy electrons to bind to the surface and cancel the electric field from the Rb adsorbates. Our results will be important for integrating Rydberg atoms into hybrid quantum systems, as fundamental probes of atom-surface interactions, and for studies of 2D electron gases bound to surfaces. PMID:27081976

  13. Metric-affine formalism of higher derivative scalar fields in cosmology

    SciTech Connect

    Li, Mingzhe; Wang, Xiulian E-mail: wangxiulian2000@yahoo.com.cn

    2012-07-01

    Higher derivative scalar field theories have received considerable attention for the potentially explanations of the initial state of the universe or the current cosmic acceleration which they might offer. They have also attracted many interests in the phenomenological studies of infrared modifications of gravity. These theories are mostly studied by the metric variational approach in which only the metric is the fundamental field to account for the gravitation. In this paper we study the higher derivative scalar fields with the metric-affine formalism where the affine connection is treated arbitrarily at the beginning. Because the higher derivative scalar fields couple to the connection directly in a covariant theory these two formalisms will lead to different results. These differences are suppressed by the powers of the Planck mass and are usually expected to have small effects. But in some cases they may cause non-negligible deviations. We show by a higher derivative dark energy model that the two formalisms lead to significantly different pictures of the future universe.

  14. Toda Systems, Cluster Characters, and Spectral Networks

    NASA Astrophysics Data System (ADS)

    Williams, Harold

    2016-11-01

    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.

  15. Electric field cancellation on quartz by Rb adsorbate-induced negative electron affinity

    NASA Astrophysics Data System (ADS)

    Shaffer, James

    2016-05-01

    We investigate the (0001) surface of single crystal quartz with a submonolayer of Rb adsorbates. Using Rydberg atom electromagnetically induced transparency, we investigate the electric fields resulting from Rb adsorbed on the quartz surface, and measure the activation energy of the Rb adsorbates. We show that the adsorbed Rb induces a negative electron affinity (NEA) on the quartz surface. The NEA surface allows low energy electrons to bind to the surface and cancel the electric field from the Rb adsorbates. Our results are important for integrating Rydberg atoms into hybrid quantum systems and the fundamental study of atom-surface interactions, as well as applications for electrons bound to a 2D surface. This work was supported by the DARPA Quasar program by a Grant through ARO (60181-PH-DRP) and the AFOSR (FA9550-12-1-0282),.

  16. Evaluation of the surface affinity of water in three biochars using fast field cycling NMR relaxometry.

    PubMed

    Bubici, Salvatore; Korb, Jean-Pierre; Kučerik, Jiří; Conte, Pellegrino

    2016-05-01

    Many soil functions depend on the interaction of water with soil. The affinity of water for soils can be altered by applying soil amendments like stone meal, manure, or biochar (a carbonaceous material obtained by pyrolysis of biomasses). In fact, the addition of hydrophobic biochar to soil may increase soil repellency, reduce water-adsorbing capacity, inhibit microbial activity, alter soil filter, buffer, storage, and transformation functions. For this reason, it is of paramount importance to monitor water affinity for biochar surface (also referred to as 'wettability') in order to better address its applications in soil systems. In this study, we propose the use of fast field cycling NMR relaxometry technique with the application of a new mathematical model for data interpretation, as a valid alternative to the traditional contact angle (CA) measurements for biochar wettability evaluation. Either NMR or CA results revealed the same wettability trend for the biochars studied here. The advantage of NMR relaxometry over CA measurements lies in the possibility to obtain at the microscopic level a variety of different information in only one shot. In fact, while CA provides only wettability evaluation, NMR relaxometry also allows achievement of the mechanisms for water molecular dynamics on biochar surface, thereby leading to the possibility to understand better, in future research, the role of biochar in increasing soil quality and plant nutrition. PMID:27062147

  17. 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.

  18. On polynomial vector fields having a given affine variety as attractive and invariant set: application to robotics

    NASA Astrophysics Data System (ADS)

    Possieri, Corrado; Tornambè, Antonio

    2015-05-01

    The main goal of this paper is to compute a class of polynomial vector fields, whose associated dynamical system has a given affine variety as attractive and invariant set, a given point in such an affine variety as invariant and attractive and another given affine variety as invariant set, solving the application of this technique in the robotic area. This objective is reached by using some tools taken from algebraic geometry. Practical examples of how these vector fields can be computed are reported. Moreover, by using these techniques, two feedback control laws, respectively, for a unicycle-like mobile robot and for a car-like mobile robot, which make them move, within the workspace, approaching to a selected algebraic curve, are given.

  19. Ultradiscretization of the theta function solution of pd Toda

    NASA Astrophysics Data System (ADS)

    Iwao, Shinsuke; Tokihiro, Tetsuji

    2007-10-01

    A periodic box-ball system (pBBS) is obtained by ultradiscretizing the periodic discrete Toda equation (pd Toda equation). We show the relation between a Young diagram of the pBBS and a spectral curve of the pd Toda equation. The formula for the fundamental cycle of the pBBS is obtained as a corollary.

  20. Toric Networks, Geometric R-Matrices and Generalized Discrete Toda Lattices

    NASA Astrophysics Data System (ADS)

    Inoue, Rei; Lam, Thomas; Pylyavskyy, Pavlo

    2016-11-01

    We use the combinatorics of toric networks and the double affine geometric R-matrix to define a three-parameter family of generalizations of the discrete Toda lattice. We construct the integrals of motion and a spectral map for this system. The family of commuting time evolutions arising from the action of the R-matrix is explicitly linearized on the Jacobian of the spectral curve. The solution to the initial value problem is constructed using Riemann theta functions.

  1. Toric Networks, Geometric R-Matrices and Generalized Discrete Toda Lattices

    NASA Astrophysics Data System (ADS)

    Inoue, Rei; Lam, Thomas; Pylyavskyy, Pavlo

    2016-08-01

    We use the combinatorics of toric networks and the double affine geometric R-matrix to define a three-parameter family of generalizations of the discrete Toda lattice. We construct the integrals of motion and a spectral map for this system. The family of commuting time evolutions arising from the action of the R-matrix is explicitly linearized on the Jacobian of the spectral curve. The solution to the initial value problem is constructed using Riemann theta functions.

  2. Prediction of SAMPL3 host-guest binding affinities: evaluating the accuracy of generalized force-fields.

    PubMed

    Muddana, Hari S; Gilson, Michael K

    2012-05-01

    We used the second-generation mining minima method (M2) to compute the binding affinities of the novel host-guest complexes in the SAMPL3 blind prediction challenge. The predictions were in poor agreement with experiment, and we conjectured that much of the error might derive from the force field, CHARMm with Vcharge charges. Repeating the calculations with other generalized force-fields led to no significant improvement, and we observed that the predicted affinities were highly sensitive to the choice of force-field. We therefore embarked on a systematic evaluation of a set of generalized force fields, based upon comparisons with PM6-DH2, a fast yet accurate semi-empirical quantum mechanics method. In particular, we compared gas-phase interaction energies and entropies for the host-guest complexes themselves, as well as for smaller chemical fragments derived from the same molecules. The mean deviations of the force field interaction energies from the quantum results were greater than 3 kcal/mol and 9 kcal/mol, for the fragments and host-guest systems respectively. We further evaluated the accuracy of force-fields for computing the vibrational entropies and found the mean errors to be greater than 4 kcal/mol. Given these errors in energy and entropy, it is not surprising in retrospect that the predicted binding affinities deviated from the experiment by several kcal/mol. These results emphasize the need for improvements in generalized force-fields and also highlight the importance of systematic evaluation of force-field parameters prior to evaluating different free-energy methods.

  3. Non-affine fields in solid-solid transformations: the structure and stability of a product droplet.

    PubMed

    Paul, Arya; Sengupta, Surajit; Rao, Madan

    2014-01-01

    We describe the microstructure, morphology, and dynamics of growth of a droplet of martensite nucleating in a parent austenite during a solid-solid transformation, using a Landau theory written in terms of both conventional affine elastic deformations and non-affine deformations. Non-affineness, φ, serves as a source of strain incompatibility and screens long-ranged elastic interactions. It is produced wherever the local stress exceeds a threshold and anneals diffusively thereafter. Using a variational calculation, we find three types of stable solution (labeled I, II, and III) for the structure of the product droplet, depending on the stress threshold and the scaled mobilities of φ parallel and perpendicular to the parent-product interface. The profile of the non-affine field φ is different in these three solutions: I is characterized by a vanishingly small φ, II admits large values of φ localized in regions of high stress within the parent-product interface, and III is a structure in which φ completely wets the parent-product interface. The width l and size W of the twins follow the relation l is proportional to √W in solution I; this relation does not hold for II or III. We obtain a dynamical phase diagram featuring these solutions, and argue that they represent specific solid-state microstructures.

  4. Wave Equations for Classical Two-Component Proca Fields in Curved Spacetimes with Torsionless Affinities

    NASA Astrophysics Data System (ADS)

    Santos Júnior, S. I.; Cardoso, J. G.

    2016-10-01

    The world formulation of the full theory of classical Proca fields in generally relativistic spacetimes is reviewed. Subsequently, the entire set of field equations is transcribed in a straightforward way into the framework of one of the Infeld-van der Waerden formalisms. Some well-known calculational techniques are then utilized for deriving the wave equations that control the propagation of the fields allowed for. It appears that no interaction couplings between such fields and electromagnetic curvatures are ultimately carried by the wave equations at issue. What results is, in effect, that the only interactions which occur in the theoretical context under consideration involve strictly Proca fields and wave functions for gravitons.

  5. Two-Component Theory of Classical Proca Fields in Curved Spacetimes with Torsionless Affinities

    NASA Astrophysics Data System (ADS)

    Santos Júnior, S. I.; Cardoso, J. G.

    2016-04-01

    The world formulation of the full theory of classical Proca fields in generally relativistic spacetimes is reviewed. Subsequently the entire set of field equations is transcribed in a straightforward way into the framework of one of the Infeld-van der Waerden formalisms. Some well-known calculational techniques are then utilized for deriving the wave equations that control the propagation of the fields allowed for. It appears that no interaction couplings between such fields and electromagnetic curvatures are ultimately carried by the wave equations at issue. What results is, in effect, that the only interactions which occur in the theoretical context under consideration involve strictly Proca fields and wave functions for gravitons.

  6. Effect of Pentacene-dielectric Affinity on Pentacene Thin Film Growth Morphology in Organic Field-effect Transistors

    SciTech Connect

    S Kim; M Jang; H Yang; C Park

    2011-12-31

    Organic field-effect transistors (OFETs) are fabricated by depositing a thin film of semiconductor on the functionalized surface of a SiO{sub 2} dielectric. The chemical and morphological structures of the interface between the semiconductor and the functionalized dielectric are critical for OFET performance. We have characterized the effect of the affinity between semiconductor and functionalized dielectric on the properties of the semiconductor-dielectric interface. The crystalline microstructure/nanostructure of the pentacene semiconductor layers, grown on a dielectric substrate that had been functionalized with either poly(4-vinyl pyridine) or polystyrene (to control hydrophobicity), and grown under a series of substrate temperatures and deposition rates, were characterized by X-ray diffraction, photoemission spectroscopy, and atomic force microscopy. By comparing the morphological features of the semiconductor thin films with the device characteristics (field-effect mobility, threshold voltage, and hysteresis) of the OFET devices, the effect of affinity-driven properties on charge modulation, charge trapping, and charge carrier transport could be described.

  7. Statistical mechanics of the Toda lattice based on soliton dynamics

    NASA Astrophysics Data System (ADS)

    Yoshida, Fumio; Sakurma, Tetsuro

    1982-05-01

    A classical theory of statistical mechanics of the Toda lattice is presented on the basis of soliton dynamics. Following the inverse spectral theory, the partition function of the Toda lattice is reconstructed from one-particle partition functions of soliton and ripple modes. Discussions are made on the contribution of these modes to the thermodynamic properties of the Toda lattice. At low temperatures, it is shown that the average number of excited solitons has the temperature dependence T13. With the comparison of our results with those from the exact theory, several problems to be worked out are pointed out in our soliton-ripple gas-mixture model.

  8. M2-brane surface operators and gauge theory dualities in Toda

    NASA Astrophysics Data System (ADS)

    Gomis, Jaume; Le Floch, Bruno

    2016-04-01

    We give a microscopic two dimensional {N} = (2, 2) gauge theory description of arbitrary M2-branes ending on N f M5-branes wrapping a punctured Riemann surface. These realize surface operators in four dimensional {N} = 2 field theories. We show that the expectation value of these surface operators on the sphere is captured by a Toda CFT correlation function in the presence of an additional degenerate vertex operator labelled by a representation {R} of SU( N f ), which also labels M2-branes ending on M5-branes. We prove that symmetries of Toda CFT correlators provide a geometric realization of dualities between two dimensional gauge theories, including {N} = (2, 2) analogues of Seiberg and Kutasov-Schwimmer dualities. As a bonus, we find new explicit conformal blocks, braiding matrices, and fusion rules in Toda CFT.

  9. 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)

  10. Phylogenetic affinities of Phobetinus to other pirate spider genera (Araneae: Mimetidae) as indicated by spinning field morphology.

    PubMed

    Townley, Mark A; Harms, Danilo; Benjamin, Suresh P

    2013-09-01

    Spinnerets from Phobetinus sagittifer and an undescribed Phobetinus species were examined by scanning electron microscopy to gain a better understanding of this genus' relationships to other genera in the family Mimetidae. Consistent with placement of Phobetinus in Mimetinae, females possessed two synapomorphies of this subfamily; enlarged cylindrical silk gland spigots with domed shafts and a single cylindrical spigot per posterior lateral spinneret (PLS). Spinning field features overall suggest Phobetinus is most closely related to Mimetus, followed by Australomimetus, then Ero. A possible synapomorphy of a clade including Mimetus and Phobetinus is a pair of modified piriform silk gland spigots on each anterior lateral spinneret of adult males located adjacent to the secondary major ampullate silk gland tartipore. These spigots were present in P. sagittifer; however, similarly positioned spigots in the undescribed species were not obviously modified (i.e., wider or with larger openings relative to the other piriform spigots). Close affinity to Mimetus was also indicated by tartipore-accommodated PLS aciniform silk glands in both Phobetinus species. These have been consistently observed in Mimetus, but not in Australomimetus or Ero. Somatic and genitalic drawings of P. sagittifer are provided to aid identification and similarities are noted between male pedipalps of Mimetus and Phobetinus.

  11. Conformal field theories from deformations of theories with Wn symmetry

    NASA Astrophysics Data System (ADS)

    Babaro, Juan Pablo; Giribet, Gaston; Ranjbar, Arash

    2016-10-01

    We construct a set of nonrational conformal field theories that consist of deformations of Toda field theory for s l (n ). In addition to preserving conformal invariance, the theories may still exhibit a remnant infinite-dimensional affine symmetry. The case n =3 is used to illustrate this phenomenon, together with further deformations that yield enhanced Kac-Moody symmetry algebras. For generic n we compute N -point correlation functions on the Riemann sphere and show that these can be expressed in terms of s l (n ) Toda field theory ((N -2 )n +2 ) -point correlation functions.

  12. Orbifold melting crystal models and reductions of Toda hierarchy

    NASA Astrophysics Data System (ADS)

    Takasaki, Kanehisa

    2015-05-01

    Orbifold generalizations of the ordinary and modified melting crystal models are introduced. They are labelled by a pair a,b of positive integers, and geometrically related to {{{Z}}a}× {{{Z}}b} orbifolds of local {C}{{{P}}1} geometry of the {O}(0)\\oplus {O}(-2) and {O}(-1)\\oplus {O}(-1) types. The partition functions have a fermionic expression in terms of charged free fermions. With the aid of shift symmetries in a fermionic realization of the quantum torus algebra, one can convert these partition functions to tau functions of the 2D Toda hierarchy. The powers {{L}a},{{\\bar{L}}-b} of the associated Lax operators turn out to take a special factorized form that defines a reduction of the 2D Toda hierarchy. The reduced integrable hierarchy for the orbifold version of the ordinary melting crystal model is the bi-graded Toda hierarchy of bi-degree (a,b). That of the orbifold version of the modified melting crystal model is the rational reduction of bi-degree (a,b). This result seems to be in accord with recent work of Brini et al on a mirror description of the genus-zero Gromov-Witten theory on a {{{Z}}a}× {{{Z}}b} orbifold of the resolved conifold.

  13. Mass transport theory for the Toda lattices, dispersive and dissipative

    NASA Astrophysics Data System (ADS)

    Horii, Zene

    2005-05-01

    To establish mass transport theory on nonlinear lattices, we formulate the Korteweg-deVries (KdV) equation and the Burgers equation using the flow variable representation so as to facilitate comparison with the Boltzmann equation and with the Cahn-Hilliard equation in classical statistical mechanics. We also study Toda lattice microdynamics using the Flaschka representation, and compare with the Liouville equation. Like the linear diffusion equation, the Boltzmann equation and the Liouville equation are to be solved for a distribution function, which is intrinsically probabilistic. Transport theory in linear systems is governed by the isotropic motions of the kinetic equations. In contrast, the KdV perturbation equation derived from the Toda lattice microdynamics expresses hydrodynamic mass transport. The KdV equation in hydrodynamics and the Burgers equation in thermodynamics do not involve a probability distribution function. The nonlinear lattices do not retain isotropy of the mass transport equations. In consequence, it is proposed that in the presence of hydrodynamic flows to the left, KdV wave propagation proceeds to the right. This basic property of the KdV system is extended to thermodynamics in the Burgers system. These features arise because linear systems are driven towards an equilibrium by molecular collisions, whereas the inhomogeneities of the nonlinear lattices are generated by the potential energy of interaction. Diffusion as expressed by the Burgers equation is governed not only by a chemical potential, but also by the Toda lattice potential energy.

  14. An(1) Toda solitons and the dressing symmetry

    NASA Astrophysics Data System (ADS)

    Belich, H.; Paunov, R.

    1997-08-01

    We present an elementary derivation of the solitonlike solutions in the An(1) Toda models which is an alternative to the previously used Hirota method. The solutions of the underlying linear problem corresponding to the N-solitons are calculated. This enables us to obtain explicit expression for the element which, by dressing group action, produces a generic soliton solution. In the particular example of monosolitons we suggest a relation to the vertex operator formalism, previously used by Olive, Turok, and Underwood. Our results can also be considered as generalization of the approach to the sine-Gordon solitons, proposed by Babelon and Bernard.

  15. 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.

  16. Low uptake affinity cultivars with biochar to tackle Cd-tainted rice--A field study over four rice seasons in Hunan, China.

    PubMed

    Chen, De; Guo, Hu; Li, Ruiyue; Li, Lianqing; Pan, Genxing; Chang, Andrew; Joseph, Stephen

    2016-01-15

    Biochar is becoming an environmentally friendly material for remediation of heavy metal contaminated soils and improving food safety. A field trial over four rice seasons was conducted to investigate the use of biochar and low Cd accumulating cultivars on Cd uptake in a heavy metal contaminated soil. Wheat straw derived biochar was applied at 0, 20 and 40 t ha(-1). Two rice cultivars with differing Cd accumulation abilities were selected in each season. The results showed that both biochar and low Cd affinity cultivars significantly reduced rice grain Cd accumulation. Biochar had no significant effect the first season but thereafter consistently reduced rice grain Cd by a maximum of 61, 86 and 57% over the next three seasons. Zn accumulation in the rice grains was not decreased by biochar application, although available soil Zn was sharply reduced (35-91%). Indica conventional rice cultivars had much lower Cd, but higher Zn and lower Cd/Zn ratios in the grain than indica hybrid cultivars. Biochar was more effective for mitigating grain Cd accumulation in low Cd affinity cultivars than in high affinity cultivars. Soil pH was sustainably increased (up to nearly 1 unit) while available Cd significantly decreased by a maximum of 85% after biochar addition. The translocation of Cd from rice roots to shoots was reduced from 20 to 80% by biochar. Low uptake affinity cultivars combined with biochar reduced late rice grain Cd concentration and Cd/Zn ratios by 69-80% and 72-80%, respectively. It indicated that the management of combining biochar and low Cd affinity cultivars should be an efficient way to remediate Cd contaminated rice paddies and reduce health risk associated with consuming rice from these soils.

  17. 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

  18. 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.

  19. 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)

  20. 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.

  1. Combinatorial expressions of the solutions to initial value problems of the discrete and ultradiscrete Toda molecules

    NASA Astrophysics Data System (ADS)

    Kamioka, Shuhei; Takagaki, Tomoaki

    2013-09-01

    Combinatorial expressions are presented of the solutions to initial value problems of the discrete and ultradiscrete Toda molecules. For the discrete Toda molecule, a subtraction-free expression of the solution is derived in terms of non-intersecting paths, for which two results in combinatorics, Flajolet’s interpretation of continued fractions and Gessel-Viennot’s lemma on determinants, are applied. By ultradiscretizing the subtraction-free expression, the solution to the ultradiscrete Toda molecule is obtained. It is finally shown that the initial value problem of the ultradiscrete Toda molecule is exactly solved in terms of shortest paths on a specific graph. The behavior of the solution is also investigated in comparison with the box-ball system.

  2. Studies in Quantum Field Theory. Final Report, July 21, 1992 - July 31, 1999

    SciTech Connect

    Caldi, Daniel G.

    2001-03-31

    Mechanisms have been investigated for chiral symmetry breaking in QED and non-abelian gauge theories using the Schwinger proper-time formalism. Multi-soliton and plane-wave solutions have been generated in affine Toda field theories. New predictions for neutrino mass generation via superfluid-type condensates in the Electroweak theory have been made. Solutions for the linear inhomogeneous bioheat equation were studied in cylindrical geometries.

  3. Toda-like (0,2) mirrors to products of projective spaces

    NASA Astrophysics Data System (ADS)

    Chen, Zhuo; Sharpe, Eric; Wu, Ruoxu

    2016-08-01

    One of the open problems in understanding (0,2) mirror symmetry concerns the construction of Toda-like Landau-Ginzburg mirrors to (0,2) theories on Fano spaces. In this paper, we begin to fill this gap by making an ansatz for (0,2) Toda-like theories mirror to (0,2) supersymmetric nonlinear sigma models on products of projective spaces, with deformations of the tangent bundle, generalizing a special case previously worked out for {{P}}^1× {{P}}^1 . We check this ansatz by matching correlation functions of the B/2-twisted Toda-like theories to correlation functions of corresponding A/2-twisted nonlinear sigma models, computed primarily using localization techniques. These (0,2) Landau-Ginzburg models admit redundancies, which can lend themselves to multiple distinct-looking representatives of the same physics, which we discuss.

  4. On Non-Topological Solutions for Planar Liouville Systems of Toda-Type

    NASA Astrophysics Data System (ADS)

    Poliakovsky, Arkady; Tarantello, Gabriella

    2016-10-01

    Motivated by the study of non-abelian Chern Simons vortices of non-topological type in Gauge Field Theory, see e.g. Gudnason (Nucl Phys B 821:151-169, 2009), Gudnason (Nucl Phys B 840:160-185, 2010) and Dunne (Lecture Notes in Physics, New Series, vol 36. Springer, Heidelberg, 1995), we analyse the solvability of the following (normalised) Liouville-type system in the presence of singular sources: (1)_τ -Δ u_1 = e^{u_1} - τ e^{u_2} - 4N π δ_0,-Δ u_2 = e^{u_2} - τ e^{u_1}, β_1 = 1/2π int_{R2} e^{u_1} {and } β_2 = 1/2π int_{R2} e^{u_2}, with {τ > 0} and {N > 0}. We identify necessary and sufficient conditions on the parameter {τ} and the "flux" pair: {(β_1, β_2),} which ensure the radial solvability of {(1)_τ.} Since for {τ=1/2,} problem {(1)_τ} reduces to the (integrable) 2 {×} 2 Toda system, in particular we recover the existence result of Lin et al. (Invent Math 190(1):169-207, 2012) and Jost and Wang (Int Math Res Not 6:277-290, 2002), concerning this case. Our method relies on a blow-up analysis for solutions of {(1)_τ}, which (even in the radial setting) takes new turns compared to the single equation case. We mention that our approach also permits handling the non-symmetric case, where in each of the two equations in {(1)_τ}, the parameter {τ} is replaced by two different parameters {τ_1 > 0} and {τ_2 > 0} respectively, and also when the second equation in {(1)_τ} includes a Dirac measure supported at the origin.

  5. 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.

  6. 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.

  7. Affine projective Osserman structures

    NASA Astrophysics Data System (ADS)

    Gilkey, P.; Nikčević, S.

    2013-08-01

    By considering the projectivized spectrum of the Jacobi operator, we introduce the concept of projective Osserman manifold in both the affine and in the pseudo-Riemannian settings. If M is an affine projective Osserman manifold, then the deformed Riemannian extension metric on the cotangent bundle is both spacelike and timelike projective Osserman. Since any rank-1-symmetric space is affine projective Osserman, this provides additional information concerning the cotangent bundle of a rank-1 Riemannian symmetric space with the deformed Riemannian extension metric. We construct other examples of affine projective Osserman manifolds where the Ricci tensor is not symmetric and thus the connection in question is not the Levi-Civita connection of any metric. If the dimension is odd, we use methods of algebraic topology to show the Jacobi operator of an affine projective Osserman manifold has only one non-zero eigenvalue and that eigenvalue is real.

  8. 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).

  9. 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.

  10. 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)

  11. Bifurcation Diagrams and Quotient Topological Spaces Under the Action of the Affine Group of a Family of Planar Quadratic Vector Fields

    NASA Astrophysics Data System (ADS)

    Cerba Diaconescu, Oxana; Schlomiuk, Dana; Vulpe, Nicolae

    In this article, we consider the class QSL4{u +vc+w^c, ∞ } of all real quadratic differential systems (dx)/(dt) = p(x, y), (dy)/(dt) = q(x, y) with gcd(p, q) = 1, having invariant lines of total multiplicity four and two complex and one real infinite singularities. We first construct compactified canonical forms for the class QSL4{u +vc+w^c, ∞ } so as to include limit points in the 12-dimensional parameter space of this class. We next construct the bifurcation diagrams for these compactified canonical forms. These diagrams contain many repetitions of phase portraits and we show that these are due to many symmetries under the group action. To retain the essence of the dynamics we finally construct the quotient spaces under the action of the group G = Aff(2, ℝ) × ℝ* of affine transformations and time homotheties and we place the phase portraits in these quotient spaces. The final diagrams retain only the necessary information to capture the dynamics under the motion in the parameter space as well as under this group action. We also present here necessary and sufficient conditions for an affine line to be invariant of multiplicity k for a quadratic system.

  12. 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.

  13. Thermostatted molecular dynamics: How to avoid the Toda demon hidden in Nose-Hoover dynamics

    SciTech Connect

    Holian, B.L.; Voter, A.F.; Ravelo, R.

    1995-09-01

    The Nose-Hoover thermostat, which is often used in the hope of modifying molecular dynamics trajectories in order to achieve canonical-ensemble averages, has hidden in it a Toda ``demon,`` which can give rise to unwanted, noncanonical undulations in the instantaneous kinetic temperature. We show how these long-lived oscillations arise from insufficient coupling of the thermostat to the atoms, and give straightforward, practical procedures for avoiding this weak-coupling pathology in isothermal molecular dynamics simulations.

  14. Understanding the basis of I50V-induced affinity decrease in HIV-1 protease via molecular dynamics simulations using polarized force field.

    PubMed

    Duan, Rui; Lazim, Raudah; Zhang, Dawei

    2015-09-30

    Human immunodeficiency virus (HIV)-1 protease is one of the most promising drug target commonly utilized to combat Acquired Immune Deficiency Syndrome (AIDS). However, with the emergence of drug resistance arising from mutations, the efficiency of protease inhibitors (PIs) as a viable treatment for AIDS has been greatly reduced. I50V mutation as one of the most significant mutations occurring in HIV-1 protease will be investigated in this study. Molecular dynamics (MD) simulation was utilized to examine the effect of I50V mutation on the binding of two PIs namely indinavir and amprenavir to HIV-1 protease. Prior to the simulations conducted, the electron density distributions of the PI and each residue in HIV-1 protease are derived by combining quantum fragmentation approach molecular fractionation with conjugate caps and Poisson-Boltzmann solvation model based on polarized protein-specific charge scheme. The atomic charges of the binding complex are subsequently fitted using delta restrained electrostatic potential (delta-RESP) method to overcome the poor charge determination of buried atom. This way, both intraprotease polarization and the polarization between protease and the PI are incorporated into partial atomic charges. Through this study, the mutation-induced affinity variations were calculated and significant agreement between experiments and MD simulations conducted was observed for both HIV-1 protease-drug complexes. In addition, the mechanism governing the decrease in the binding affinity of PI in the presence of I50V mutation was also explored to provide insights pertaining to the design of the next generation of anti-HIV drugs. PMID:26198456

  15. 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.

  16. Darboux transformations of lower degree for two-dimensional C(1)l and D(2)l+1 Toda equations

    NASA Astrophysics Data System (ADS)

    Zhou, Zi-Xiang

    2008-08-01

    For the two-dimensional Toda equations corresponding to the Kac-Moody algebras C(1)l and D(2)l+1, the Darboux transformations with a special choice of spectral parameters are constructed so that the degree of these Darboux transformations is half of that for usual Darboux transformations and the derived solutions become simpler. These Darboux transformations for a Lax pair are constructed from real solutions of itself or a slightly different Lax pair corresponding to the same Toda equation, depending on the parity of l. Exact solutions of these Toda equations are presented by simplifying the results derived from the Darboux transformations.

  17. Affinity driven social networks

    NASA Astrophysics Data System (ADS)

    Ruyú, B.; Kuperman, M. N.

    2007-04-01

    In this work we present a model for evolving networks, where the driven force is related to the social affinity between individuals of a population. In the model, a set of individuals initially arranged on a regular ordered network and thus linked with their closest neighbors are allowed to rearrange their connections according to a dynamics closely related to that of the stable marriage problem. We show that the behavior of some topological properties of the resulting networks follows a non trivial pattern.

  18. The Toda System and Clustering Interfaces in the Allen Cahn equation

    NASA Astrophysics Data System (ADS)

    Del Pino, Manuel; Kowalczyk, Michał; Wei, Juncheng

    2008-10-01

    We consider the Allen Cahn equation {\\varepsilon2Δ u + (1-u^2)u = 0} in a bounded, smooth domain Ω in {mathbb{R}^2} , under zero Neumann boundary conditions, where {\\varepsilon > 0} is a small parameter. Let Γ0 be a segment contained in Ω, connecting orthogonally the boundary. Under certain nondegeneracy and nonminimality assumptions for Γ0, satisfied for instance by the short axis in an ellipse, we construct, for any given N ≥ 1, a solution exhibiting N transition layers whose mutual distances are {O(\\varepsilon|log\\varepsilon|)} and which collapse onto Γ0 as {\\varepsilonto 0} . Asymptotic location of these interfaces is governed by a Toda-type system and yields in the limit broken lines with an angle at a common height and at main order cutting orthogonally the boundary.

  19. 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

  20. BC(50): a generalized, unifying affinity descriptor.

    PubMed

    Vacca, Alberto; Francesconi, Oscar; Roelens, Stefano

    2012-12-01

    Assessing binding affinities is an unavoidable step that we come across any time interactions between binding species are investigated. A quantitative evaluation of binding affinities relies on the determination of binding constants but, whilst the binding constant fully defines the affinity of a reagent for a ligand when only one complex species is formed, the same is not true when the interacting partners form more than one complex of different stoichiometry, because all complexes contribute to the overall binding affinity. Unfortunately, this situation is the rule rather than the exception in chemical systems, but a generally accepted solution for this issue has not yet been settled. In this Personal Account, we describe the evolution, from the initial idea to a fully developed stage, of a binding descriptor that has been developed with the aim of filling this gap, thereby providing scientists in all fields of chemistry with a unifying tool for the assessment of binding affinities based on the knowledge of the binding constants in systems that involve any number of complex species.

  1. Long-time asymptotics of the periodic Toda lattice under short-range perturbations

    NASA Astrophysics Data System (ADS)

    Kamvissis, Spyridon; Teschl, Gerald

    2012-07-01

    We compute the long-time asymptotics of periodic (and slightly more generally of algebro-geometric finite-gap) solutions of the doubly infinite Toda lattice under a short-range perturbation. In particular, we prove that the perturbed lattice asymptotically approaches a modulated lattice. More precisely, let g be the genus of the hyperelliptic curve associated with the unperturbed solution. We show that, apart from the phenomenon of solitons travelling in a quasi-periodic background, the n/t-pane contains g + 2 areas where the perturbed solution is close to a finite-gap solution on the same isospectral torus. In between there are g + 1 regions where the perturbed solution is asymptotically close to a modulated lattice which undergoes a continuous phase transition (in the Jacobian variety) and which interpolates between these isospectral solutions. In the special case of the free lattice (g = 0), the isospectral torus consists of just one point and we recover the known result. Both the solutions in the isospectral torus and the phase transition are explicitly characterized in terms of Abelian integrals on the underlying hyperelliptic curve. Our method relies on the equivalence of the inverse spectral problem to a vector Riemann-Hilbert problem defined on the hyperelliptic curve and generalizes the so-called nonlinear stationary phase/steepest descent method for Riemann-Hilbert problem deformations to Riemann surfaces.

  2. 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.

  3. 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.

  4. 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

  5. Overview of affinity biosensors in food analysis.

    PubMed

    Patel, Pradip D

    2006-01-01

    The 4 major driving forces that are expected to lead to increased use of affinity biosensors that meet crucial industrial test specifications, e.g., fast, reliable, cost-effective, and use of low-skilled personnel, are (1) strict legislative framework, e.g., recent changes proposed to the European food safety and hygiene legislation, EC No. 178/2002; (2) industrial shift from quality control to quality assurance procedures, e.g., Hazard Analysis Critical Control Point, ensuring effective positioning in the global competitive trade; (3) just-in-time production resulting in 'right' product every time; and (4) consumer demand for safe and wholesome products. The affinity biosensors field has expanded significantly over the past decade, with a projected global biosensors market growth from $6.1 billion in 2004 to $8.2 billion in 2009, representing major industrial sectors (e.g., Pharma, Medicare, and Food). This brief review is targeted to affinity biosensors developed for the food industry and includes research and development leading to biosensors for microbiological and chemical analytes of industrial concern, commercial biosensors products on the market, and examples of future prospects in this diagnostic field.

  6. Overview of affinity biosensors in food analysis.

    PubMed

    Patel, Pradip D

    2006-01-01

    The 4 major driving forces that are expected to lead to increased use of affinity biosensors that meet crucial industrial test specifications, e.g., fast, reliable, cost-effective, and use of low-skilled personnel, are (1) strict legislative framework, e.g., recent changes proposed to the European food safety and hygiene legislation, EC No. 178/2002; (2) industrial shift from quality control to quality assurance procedures, e.g., Hazard Analysis Critical Control Point, ensuring effective positioning in the global competitive trade; (3) just-in-time production resulting in 'right' product every time; and (4) consumer demand for safe and wholesome products. The affinity biosensors field has expanded significantly over the past decade, with a projected global biosensors market growth from $6.1 billion in 2004 to $8.2 billion in 2009, representing major industrial sectors (e.g., Pharma, Medicare, and Food). This brief review is targeted to affinity biosensors developed for the food industry and includes research and development leading to biosensors for microbiological and chemical analytes of industrial concern, commercial biosensors products on the market, and examples of future prospects in this diagnostic field. PMID:16792079

  7. 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.

  8. Cesium cation affinities and basicities

    NASA Astrophysics Data System (ADS)

    Gal, Jean-François; Maria, Pierre-Charles; Massi, Lionel; Mayeux, Charly; Burk, Peeter; Tammiku-Taul, Jaana

    2007-11-01

    This review focuses on the quantitative data related to cesium cation interaction with neutral or negatively charged ligands. The techniques used for measuring the cesium cation affinity (enthalpies, CCA), and cesium cation basicities (Gibbs free energies, CCB) are briefly described. The quantum chemical calculations methods that were specifically designed for the determination of cesium cation adduct structures and the energetic aspects of the interaction are discussed. The experimental results, obtained essentially from mass spectrometry techniques, and complemented by thermochemical data, are tabulated and commented. In particular, the correlations between cesium cation affinities and lithium cation affinities for the various kinds of ligands (rare gases, polyatomic neutral molecules, among them aromatic compounds and negative ions) serve as a basis for the interpretation of the diverse electrostatic modes of interaction. A brief account of some recent analytical applications of ion/molecule reactions with Cs+, as well as other cationization approaches by Cs+, is given.

  9. "Clickable" agarose for affinity chromatography.

    PubMed

    Punna, Sreenivas; Kaltgrad, Eiton; Finn, M G

    2005-01-01

    Successful purification of biological molecules by affinity chromatography requires the attachment of desired ligands to biocompatible chromatographic supports. The Cu(I)-catalyzed cycloaddition of azides and alkynes-the premier example of "click chemistry"-is an efficient way to make covalent connections among diverse molecules and materials. Both azide and alkyne units are highly selective in their reactivity, being inert to most chemical functionalities and stable to wide ranges of solvent, temperature, and pH. We show that agarose beads bearing alkyne and azide groups can be easily made and are practical precursors to functionalized agarose materials for affinity chromatography.

  10. 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

  11. 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…

  12. 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.…

  13. The metrizability problem for Lorentz-invariant affine connections

    NASA Astrophysics Data System (ADS)

    Urban, Zbyněk; Volná, Jana

    2016-07-01

    The invariant metrizability problem for affine connections on a manifold, formulated by Tanaka and Krupka for connected Lie groups actions, is considered in the particular cases of Lorentz and Poincaré (inhomogeneous Lorentz) groups. Conditions under which an affine connection on the open submanifold ℝ × (ℝ3\\{(0, 0, 0)}) of the Euclidean space ℝ4 coincides with the Levi-Civita connection of some SO(3, 1), respectively (ℝ4 × sSO(3, 1))-invariant metric field are studied. We give complete description of metrizable Lorentz-invariant connections. Explicit solutions (metric fields) of the invariant metrizability equations are found and their properties are discussed.

  14. 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

  15. 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.

  16. 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.

  17. Two-parameter twisted quantum affine algebras

    NASA Astrophysics Data System (ADS)

    Jing, Naihuan; Zhang, Honglian

    2016-09-01

    We establish Drinfeld realization for the two-parameter twisted quantum affine algebras using a new method. The Hopf algebra structure for Drinfeld generators is given for both untwisted and twisted two-parameter quantum affine algebras, which include the quantum affine algebras as special cases.

  18. 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.

  19. 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

  20. 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

  1. 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.

  2. 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

  3. 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.

  4. 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

  5. Engineering antibody affinity and specificity.

    PubMed

    Webster, D M; Roberts, S; Cheetham, J C; Griest, R; Rees, A R

    1988-01-01

    A combination of ab initio calculations, "knowledge-based prediction", molecular graphics and site-directed mutagenesis has enabled us to probe the molecular details of antibody:antigen recognition and binding and to alter the affinity and specificity of an antibody for its antigen. The significance of electrostatic hydrogen bonding, hydrophilic/hydrophobic patch matching and van der Waals interactions as well as CDR:CDR interactions are discussed in relation to the results of site-directed mutagenesis experiments on the anti-lysozyme antibody Gloop2. The ability to generate reconstructed antibodies, chimeric antibodies, catalytic antibodies and the use of modelled antibodies for the design of drugs is discussed. PMID:3209295

  6. Proton affinities of hydrated molecules

    NASA Astrophysics Data System (ADS)

    Valadbeigi, Younes

    2016-09-01

    Proton affinities (PA) of non-hydrated, M, and hydrated forms, M(H2O)1,2,3, of 20 organic molecules including alcohols, ethers, aldehydes, ketones and amines were calculated by the B3LYP/6-311++G(d,p) method. For homogeneous families, linear correlations were observed between PAs of the M(H2O)1,2,3 and the PAs of the non-hydrated molecules. Also, the absolute values of the hydration enthalpies of the protonated molecules decreased linearly with the PAs. The correlation functions predicted that for an amine with PA < 1100 kJ/mol the PA(M(H2O)) is larger than the corresponding PA, while for an amine with PA > 1100 kJ/mol the PA(M(H2O)) is smaller than the PA.

  7. 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.

  8. Non-affine deformations in polymer hydrogels

    PubMed Central

    Wen, Qi; Basu, Anindita; Janmey, Paul A.; Yodh, A. G.

    2012-01-01

    Most theories of soft matter elasticity assume that the local strain in a sample after deformation is identical everywhere and equal to the macroscopic strain, or equivalently that the deformation is affine. We discuss the elasticity of hydrogels of crosslinked polymers with special attention to affine and non-affine theories of elasticity. Experimental procedures to measure non-affine deformations are also described. Entropic theories, which account for gel elasticity based on stretching out individual polymer chains, predict affine deformations. In contrast, simulations of network deformation that result in bending of the stiff constituent filaments generally predict non-affine behavior. Results from experiments show significant non-affine deformation in hydrogels even when they are formed by flexible polymers for which bending would appear to be negligible compared to stretching. However, this finding is not necessarily an experimental proof of the non-affine model for elasticity. We emphasize the insights gained from experiments using confocal rheoscope and show that, in addition to filament bending, sample micro-inhomogeneity can be a significant alternative source of non-affine deformation. PMID:23002395

  9. 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.

  10. 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.

  11. pp-waves with torsion and metric-affine gravity

    NASA Astrophysics Data System (ADS)

    Pasic, Vedad; Vassiliev, Dmitri

    2005-10-01

    A classical pp-wave is a four-dimensional Lorentzian spacetime which admits a nonvanishing parallel spinor field; here the connection is assumed to be Levi-Civita. We generalize this definition to metric compatible spacetimes with torsion and describe basic properties of such spacetimes. We use our generalized pp-waves for constructing new explicit vacuum solutions of quadratic metric-affine gravity.

  12. 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

  13. 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.

  14. 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.

  15. Parafermionic representation of the affine /sl(21C) algebra at fractional level

    NASA Astrophysics Data System (ADS)

    Bowcock, P.; Hayes, M.; Taormina, A.

    1999-12-01

    The four fermionic currents of the affine superalgebra /sl(21C) at fractional level k=1/u-1,u∈N are shown to be realised in terms of a free scalar field, an /sl(2C) doublet field and primary fields of the parafermionic algebra Zu-1.

  16. 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

  17. 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...

  18. Visualizing Antibody Affinity Maturation in Germinal Centers

    PubMed Central

    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.

    2016-01-01

    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 non-immunodominant specificities must be elicited, as is the case for HIV-1 and influenza. PMID:26912368

  19. 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

  20. Designing Chaotic Systems by Piecewise Affine Systems

    NASA Astrophysics Data System (ADS)

    Wu, Tiantian; Li, Qingdu; Yang, Xiao-Song

    Based on mathematical analysis, this paper provides a methodology to ensure the existence of homoclinic orbits in a class of three-dimensional piecewise affine systems. In addition, two chaotic generators are provided to illustrate the effectiveness of the method.

  1. Affinity Electrophoresis Using Ligands Attached To Polymers

    NASA Technical Reports Server (NTRS)

    Van Alstine, James M.; Snyder, Robert S.; Harris, J. M.; Brooks, D. E.

    1990-01-01

    In new technique, reduction of electrophoretic mobilities by addition of polyethylene glycol to ligands increases electrophoretic separabilities. In immuno-affinity electrophoresis, modification of ligands extends specificity of electrophoretic separation to particles having surface electric-charge structures otherwise making them electrophoretically inseparable. Modification of antibodies by polyethylene glycol greatly reduces ability to aggregate while enhancing ability to affect electrophoretic mobilities of cells. In hydrophobic-affinity electrophoresis, addition of polyethylene glycol reduces tendency toward aggregation of cells or macromolecules.

  2. Higher Sugawara Operators for the Quantum Affine Algebras of Type A

    NASA Astrophysics Data System (ADS)

    Frappat, Luc; Jing, Naihuan; Molev, Alexander; Ragoucy, Eric

    2016-07-01

    We give explicit formulas for the elements of the center of the completed quantum affine algebra in type A at the critical level that are associated with the fundamental representations. We calculate the images of these elements under a Harish-Chandra-type homomorphism. These images coincide with those in the free field realization of the quantum affine algebra and reproduce generators of the q-deformed classical {{mathcal W}}-algebra of Frenkel and Reshetikhin.

  3. 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

  4. 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.

  5. 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

  6. 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

  7. Affine.m—Mathematica package for computations in representation theory of finite-dimensional and affine Lie algebras

    NASA Astrophysics Data System (ADS)

    Nazarov, Anton

    2012-11-01

    In this paper we present Affine.m-a program for computations in representation theory of finite-dimensional and affine Lie algebras and describe implemented algorithms. The algorithms are based on the properties of weights and Weyl symmetry. Computation of weight multiplicities in irreducible and Verma modules, branching of representations and tensor product decomposition are the most important problems for us. These problems have numerous applications in physics and we provide some examples of these applications. The program is implemented in the popular computer algebra system Mathematica and works with finite-dimensional and affine Lie algebras. Catalogue identifier: AENA_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/AENB_v1_0.html Program obtainable from: CPC Program Library, Queen’s University, Belfast, UK Licensing provisions: Standard CPC licence, http://cpc.cs.qub.ac.uk/licence/licence.html No. of lines in distributed program, including test data, etc.: 24 844 No. of bytes in distributed program, including test data, etc.: 1 045 908 Distribution format: tar.gz Programming language: Mathematica. Computer: i386-i686, x86_64. Operating system: Linux, Windows, Mac OS, Solaris. RAM: 5-500 Mb Classification: 4.2, 5. Nature of problem: Representation theory of finite-dimensional Lie algebras has many applications in different branches of physics, including elementary particle physics, molecular physics, nuclear physics. Representations of affine Lie algebras appear in string theories and two-dimensional conformal field theory used for the description of critical phenomena in two-dimensional systems. Also Lie symmetries play a major role in a study of quantum integrable systems. Solution method: We work with weights and roots of finite-dimensional and affine Lie algebras and use Weyl symmetry extensively. Central problems which are the computations of weight multiplicities, branching and fusion coefficients are solved using one general recurrent

  8. 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,…

  9. Stepparents' Affinity-Seeking and Affinity-Maintaining Strategies with Stepchildren.

    ERIC Educational Resources Information Center

    Ganong, Lawrence; Coleman, Marilyn; Fine, Mark; Martin, Patricia

    1999-01-01

    Examines the strategies that stepparents use to develop and maintain affinity with stepchildren and the effects that these strategies have on the development of stepparent-stepchildren relationships. Thirty-one affinity-seeking strategies are identified. Results show that dyadic activities worked best, but it is important that stepchildren…

  10. Affinity chromatography of bacterial lactate dehydrogenases.

    PubMed Central

    Kelly, N; Delaney, M; O'Carra, P

    1978-01-01

    The affinity system used was the immobilized oxamate derivative previously used to purify mammalian lactate dehydrogenases. The bacterial dehydrogenases specific for the L-stereoisomer of lactate behaved in the same way as the mammalian enzymes, binding strongly in the presence of NADH. The D-lactate-specific enzymes, however, did not show any biospecific affinity for this gel. The L-specific enzymes could be purified to homogeneity in one affinity-chromatographic step. The D-specific enzymes could be efficiently separated from the L-specific ones and could then be further purified on an immobilized NAD derivative. The mechanism of activation of the lactate dehydrogenase from Streptococcus faecalis by fructose 1,6-bisphosphate was investigated by using the immobilized oxamate gel. PMID:666726

  11. Affinity chromatography of bacterial lactate dehydrogenases.

    PubMed

    Kelly, N; Delaney, M; O'Carra, P

    1978-06-01

    The affinity system used was the immobilized oxamate derivative previously used to purify mammalian lactate dehydrogenases. The bacterial dehydrogenases specific for the L-stereoisomer of lactate behaved in the same way as the mammalian enzymes, binding strongly in the presence of NADH. The D-lactate-specific enzymes, however, did not show any biospecific affinity for this gel. The L-specific enzymes could be purified to homogeneity in one affinity-chromatographic step. The D-specific enzymes could be efficiently separated from the L-specific ones and could then be further purified on an immobilized NAD derivative. The mechanism of activation of the lactate dehydrogenase from Streptococcus faecalis by fructose 1,6-bisphosphate was investigated by using the immobilized oxamate gel. PMID:666726

  12. Modified gravity in three dimensional metric-affine scenarios

    NASA Astrophysics Data System (ADS)

    Bambi, Cosimo; Ghasemi-Nodehi, M.; Rubiera-Garcia, D.

    2015-08-01

    We consider metric-affine scenarios where a modified gravitational action is sourced by electrovacuum fields in a three dimensional space-time. We first study the case of f (R ) theories, finding deviations near the center as compared to the solutions of general relativity. We then consider Born-Infeld gravity, which has raised a lot of interest in the last few years regarding its applications in astrophysics and cosmology, and show that new features always arise at a finite distance from the center. Several properties of the resulting space-times, in particular in presence of a cosmological constant term, are discussed.

  13. European and international collaboration in affinity proteomics.

    PubMed

    Stoevesandt, Oda; Taussig, Michael J

    2012-06-15

    In affinity proteomics, specific protein-binding molecules (a.k.a. binders), principally antibodies, are applied as reagents in proteome analysis. In recent years, advances in binder technologies have created the potential for an unprecedented view on protein expression and distribution patterns in plasma, cells and tissues and increasingly on protein function. Particular strengths of affinity proteomics methods include detecting proteins in their natural environments of cell or tissue, high sensitivity and selectivity for detection of low abundance proteins and exploiting binding actions such as functional interference in living cells. To maximise the use and impact of affinity reagents, it will be essential to create comprehensive, standardised binder collections. With this in mind, the EU FP7 programme AFFINOMICS (http://www.affinomics.org), together with the preceding EU programmes ProteomeBinders and AffinityProteome, aims to extend affinity proteomics research by generating a large-scale resource of validated protein-binding molecules for characterisation of the human proteome. Activity is directed at producing binders to about 1000 protein targets, primarily in signal transduction and cancer, by establishing a high throughput, coordinated production pipeline. An important aspect of AFFINOMICS is the development of highly efficient recombinant selection methods, based on phage, cell and ribosome display, capable of producing high quality binders at greater throughput and lower cost than hitherto. The programme also involves development of innovative and sensitive technologies for specific detection of target proteins and their interactions, and deployment of binders in proteomics studies of clinical relevance. The need for such binder generation programmes is now recognised internationally, with parallel initiatives in the USA for cancer (NCI) and transcription factors (NIH) and within the Human Proteome Organisation (HUPO). The papers in this volume of New

  14. 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

  15. Affine Invariant Character Recognition by Progressive Removing

    NASA Astrophysics Data System (ADS)

    Iwamura, Masakazu; Horimatsu, Akira; Niwa, Ryo; Kise, Koichi; Uchida, Seiichi; Omachi, Shinichiro

    Recognizing characters in scene images suffering from perspective distortion is a challenge. Although there are some methods to overcome this difficulty, they are time-consuming. In this paper, we propose a set of affine invariant features and a new recognition scheme called “progressive removing” that can help reduce the processing time. Progressive removing gradually removes less feasible categories and skew angles by using multiple classifiers. We observed that progressive removing and the use of the affine invariant features reduced the processing time by about 60% in comparison to a trivial one without decreasing the recognition rate.

  16. 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.

  17. 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-}.

  18. 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.

  19. Modern affinity reagents: Recombinant antibodies and aptamers.

    PubMed

    Groff, Katherine; Brown, Jeffrey; Clippinger, Amy J

    2015-12-01

    Affinity reagents are essential tools in both basic and applied research; however, there is a growing concern about the reproducibility of animal-derived monoclonal antibodies. The need for higher quality affinity reagents has prompted the development of methods that provide scientific, economic, and time-saving advantages and do not require the use of animals. This review describes two types of affinity reagents, recombinant antibodies and aptamers, which are non-animal technologies that can replace the use of animal-derived monoclonal antibodies. Recombinant antibodies are protein-based reagents, while aptamers are nucleic-acid-based. In light of the scientific advantages of these technologies, this review also discusses ways to gain momentum in the use of modern affinity reagents, including an update to the 1999 National Academy of Sciences monoclonal antibody production report and federal incentives for recombinant antibody and aptamer efforts. In the long-term, these efforts have the potential to improve the overall quality and decrease the cost of scientific research.

  20. 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…

  1. Two bradykinin binding sites with picomolar affinities

    SciTech Connect

    Manning, D.C.; Vavrek, R.; Stewart, J.M.; Snyder, S.H.

    1986-05-01

    Bradykinin (BK) and related peptides exert a wide range of effects on several organ systems. We have attempted to sort out these effects by studying the binding interaction of (/sup 3/H)BK at the membrane level with in vitro receptor binding techniques. High specific activity (/sup 3/H)BK and an enzyme inhibitor cocktail has enabled us to label two BK binding sites with different affinity and peptide specificity in several guinea-pig tissues. In the guinea-pig ileum the high-affinity site has an equilibrium dissociation constant (Kd) for (/sup 3/H)BK of 13 pM and a maximal number of binding sites of 8.3 pmol/g of tissue wet weight. The low-affinity guinea-pig ileum site displays a Kd of 910 pM, a maximum number of binding sites of 14 pmol/g of tissue wet weight and shows a greater selectivity for BK analogs over Lysyl-BK analogs. Two similar sites can also be discriminated in kidney and heart. The potencies of a series of BK analogs at the high-affinity guinea-pig ileum site correlate well with their potencies in contracting ileal smooth muscle. The binding of (/sup 3/H)BK in the guinea-pig ileum is inhibited by physiological concentrations of monovalent and divalent cations.

  2. Affine Metrics and Algebroid Structures: Application to General Relativity and Unification

    NASA Astrophysics Data System (ADS)

    Elyasi, Naiereh; Boroojerdian, Naser

    2012-10-01

    Affine metrics and their associated algebroid bundle are developed. These structures are applied to the general relativity and provide a mathematical structure for unification of gravity and electromagnetism. The final result is a field equation on the associated algebroid bundle that is similar to Einstein field equation but contains Einstein field equation and Maxwell equations simultaneously and contains a new equation that may have new results.

  3. Quantification of the affinities and kinetics of protein interactions using silicon nanowire biosensors

    PubMed Central

    Duan, Xuexin; Li, Yue; Rajan, Nitin K.; Routenberg, David A.; Modis, Yorgo; Reed, Mark A.

    2014-01-01

    Monitoring the binding affinities and kinetics of protein interactions is important in clinical diagnostics and drug development because such information is used to identify new therapeutic candidates. Surface plasmon resonance is at present the standard method used for such analysis, but this is limited by low sensitivity and low-throughput analysis. Here, we show that silicon nanowire field-effect transistors can be used as biosensors to measure protein–ligand binding affinities and kinetics with sensitivities down to femtomolar concentrations. Based on this sensing mechanism, we develop an analytical model to calibrate the sensor response and quantify the molecular binding affinities of two representative protein–ligand binding pairs. The rate constant of the association and dissociation of the protein–ligand pair is determined by monitoring the reaction kinetics, demonstrating that silicon nanowire field-effect transistors can be readily used as high-throughput biosensors to quantify protein interactions. PMID:22635097

  4. 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

  5. Engineered affinity proteins for tumour-targeting applications.

    PubMed

    Friedman, Mikaela; Ståhl, Stefan

    2009-05-01

    Targeting of tumour-associated antigens is an expanding treatment modality in clinical oncology as an alternative to, or in combination with, conventional treatments, such as chemotherapy, external-radiation therapy and surgery. Targeting of antigens that are unique or more highly expressed in tumours than in normal tissues can be used to increase the specificity and reduce the cytotoxic effect on normal tissues. Several targeting agents have been studied for clinical use, where monoclonal antibodies have been the ones most widely used. More than 20 monoclonal antibodies are approved for therapy today and the largest field is oncology. Advances in genetic engineering and in vitro selection technology has enabled the feasible high-throughput generation of monoclonal antibodies, antibody derivatives [e.g. scFvs, Fab molecules, dAbs (single-domain antibodies), diabodies and minibodies] and more recently also non-immunoglobulin scaffold proteins. Several of these affinity proteins have been investigated for both in vivo diagnostics and therapy. Affinity proteins in tumour-targeted therapy can affect tumour progression by altering signal transduction or by delivering a payload of toxin, drug or radionuclide. The ErbB receptor family has been extensively studied as biomarkers in tumour targeting, primarily for therapy using monoclonal antibodies. Two receptors in the ErbB family, EGFR (epidermal growth factor receptor) and HER2 (epidermal growth factor receptor 2), are overexpressed in various malignancies and associated with poor patient prognosis and are therefore interesting targets for solid tumours. In the present review, strategies are described for tumour targeting of solid tumours using affinity proteins to deliver radionuclides, either for molecular imaging or radiotherapy. Antibodies, antibody derivatives and non-immunoglobulin scaffold proteins are discussed with a certain focus on the affibody (Affibody) molecule. PMID:19341363

  6. 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.

  7. 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.

  8. Artificial Affinity Proteins as Ligands of Immunoglobulins

    PubMed Central

    Mouratou, Barbara; Béhar, Ghislaine; Pecorari, Frédéric

    2015-01-01

    A number of natural proteins are known to have affinity and specificity for immunoglobulins. Some of them are widely used as reagents for detection or capture applications, such as Protein G and Protein A. However, these natural proteins have a defined spectrum of recognition that may not fit specific needs. With the development of combinatorial protein engineering and selection techniques, it has become possible to design artificial affinity proteins with the desired properties. These proteins, termed alternative scaffold proteins, are most often chosen for their stability, ease of engineering and cost-efficient recombinant production in bacteria. In this review, we focus on alternative scaffold proteins for which immunoglobulin binders have been identified and characterized. PMID:25647098

  9. Permeability of self-affine rough fractures

    PubMed

    Drazer; Koplik

    2000-12-01

    The permeability of two-dimensional fractures with self-affine fractal roughness is studied via analytic arguments and numerical simulations. The limit where the roughness amplitude is small compared with average fracture aperture is analyzed by a perturbation method, while in the opposite case of narrow aperture, we use heuristic arguments based on lubrication theory. Numerical simulations, using the lattice Boltzmann method, are used to examine the complete range of aperture sizes, and confirm the analytic arguments. PMID:11138092

  10. A MEMS Dielectric Affinity Glucose Biosensor

    PubMed Central

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

    2013-01-01

    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

  11. 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

  12. Trematode hemoglobins show exceptionally high oxygen affinity.

    PubMed

    Kiger, L; Rashid, A K; Griffon, N; Haque, M; Moens, L; Gibson, Q H; Poyart, C; Marden, M C

    1998-08-01

    Ligand binding studies were made with hemoglobin (Hb) isolated from trematode species Gastrothylax crumenifer (Gc), Paramphistomum epiclitum (Pe), Explanatum explanatum (Ee), parasitic worms of water buffalo Bubalus bubalis, and Isoparorchis hypselobagri (Ih) parasitic in the catfish Wallago attu. The kinetics of oxygen and carbon monoxide binding show very fast association rates. Whereas oxygen can be displaced on a millisecond time scale from human Hb at 25 degrees C, the dissociation of oxygen from trematode Hb may require a few seconds to over 20 s (for Hb Pe). Carbon monoxide dissociation is faster, however, than for other monomeric hemoglobins or myoglobins. Trematode hemoglobins also show a reduced rate of autoxidation; the oxy form is not readily oxidized by potassium ferricyanide, indicating that only the deoxy form reacts rapidly with this oxidizing agent. Unlike most vertebrate Hbs, the trematodes have a tyrosine residue at position E7 instead of the usual distal histidine. As for Hb Ascaris, which also displays a high oxygen affinity, the trematodes have a tyrosine in position B10; two H-bonds to the oxygen molecule are thought to be responsible for the very high oxygen affinity. The trematode hemoglobins display a combination of high association rates and very low dissociation rates, resulting in some of the highest oxygen affinities ever observed.

  13. Informe sobre el avance del proyecto de determinaciones de V sen i para todas las estrellas B más brillantes que 6.5 mag.

    NASA Astrophysics Data System (ADS)

    Abt, H.; Levato, H.

    Se ha finalizado con la determinación de V sen i para todas las estrellas B más brillantes que 6.5 mag y al norte de -30 grados de declinación. Esta muestra involucra casi 1000 estrellas. Las observaciones fueron realizadas desde KPNO. Mientras tanto, se ha completado desde CASLEO la observación de 350 estrellas B más brillantes que 6.5 mag. al sur de -30 grados. Además de la determinación de la rotación axial se determinan, en los casos posibles, los anchos equivalentes de las líneas 4471 Ådel He I y 4481 Ådel Mg II.

  14. Latest European coelacanth shows Gondwanan affinities.

    PubMed

    Cavin, Lionel; Forey, Peter L; Buffetaut, Eric; Tong, Haiyan

    2005-06-22

    The last European fossil occurrence of a coelacanth is from the Mid-Cretaceous of the English Chalk (Turonian, 90 million years ago). Here, we report the discovery of a coelacanth from Late Cretaceous non-marine rocks in southern France. It consists of a left angular bone showing structures that imply close phylogenetic affinities with some extinct Mawsoniidae. The closest relatives are otherwise known from Cretaceous continental deposits of southern continents and suggest that the dispersal of freshwater organisms from Africa to Europe occurred in the Late Cretaceous.

  15. 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.

  16. Measuring an antibody affinity distribution molecule by molecule.

    PubMed

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

    2008-11-15

    Single molecule fluorescence microscopy was used to observe the binding and unbinding of hapten decorated quantum dots to 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.

  17. 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.

  18. 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.

  19. Advances in affinity ligand-functionalized nanomaterials for biomagnetic separation.

    PubMed

    Fields, Conor; Li, Peng; O'Mahony, James J; Lee, Gil U

    2016-01-01

    The downstream processing of proteins remains the most significant cost in protein production, and is largely attributed to rigorous chromatographic purification protocols, where the stringency of purity for biopharmaceutical products sometimes exceeds 99%. With an ever burgeoning biotechnology market, there is a constant demand for alternative purification methodologies, to ameliorate the dependence on chromatography, while still adhering to regulatory concerns over product purity and safety. In this article, we present an up-to-date view of bioseparation, with emphasis on magnetic separation and its potential application in the field. Additionally, we discuss the economic and performance benefits of synthetic ligands, in the form of peptides and miniaturized antibody fragments, compared to full-length antibodies. We propose that adoption of synthetic affinity ligands coupled with magnetic adsorbents, will play an important role in enabling sustainable bioprocessing in the future.

  20. 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.

  1. Cucurbiturils: from synthesis to high-affinity binding and catalysis.

    PubMed

    Assaf, Khaleel I; Nau, Werner M

    2015-01-21

    In the wide area of supramolecular chemistry, cucurbit[n]urils (CBn) present themselves as a young family of molecular containers, able to form stable complexes with various guests, including drug molecules, amino acids and peptides, saccharides, dyes, hydrocarbons, perfluorinated hydrocarbons, and even high molecular weight guests such as proteins (e.g., human insulin). Since the discovery of the first CBn, CB6, the field has seen tremendous growth with respect to the synthesis of new homologues and derivatives, the discovery of record binding affinities of guest molecules in their hydrophobic cavity, and associated applications ranging from sensing to drug delivery. In this review, we discuss in detail the fundamental properties of CBn homologues and their cyclic derivatives with a focus on their synthesis and their applications in catalysis.

  2. 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.

  3. Low Affinity GPCRs for Metabolic Intermediates: Challenges for Pharmacologists

    PubMed Central

    Smith, Nicola J.

    2012-01-01

    The discovery that a number of metabolites and metabolic intermediates can act through G protein-coupled receptors has attracted great interest in the field and has led to new therapeutic targets for diseases such as hypertension, type 2 diabetes, inflammation, and metabolic syndrome. However, the low apparent affinity of these ligands for their cognate receptors poses a number of challenges for pharmacologists interested in investigating receptor structure, function or physiology. Furthermore, the endogenous ligands matched to their receptors have other, well established metabolic roles and thus selectivity is difficult to achieve. This review discusses some of the issues researchers face when working with these receptors and highlights the ways in which a number of these obstacles have been overcome. PMID:22649402

  4. Advances in affinity ligand-functionalized nanomaterials for biomagnetic separation.

    PubMed

    Fields, Conor; Li, Peng; O'Mahony, James J; Lee, Gil U

    2016-01-01

    The downstream processing of proteins remains the most significant cost in protein production, and is largely attributed to rigorous chromatographic purification protocols, where the stringency of purity for biopharmaceutical products sometimes exceeds 99%. With an ever burgeoning biotechnology market, there is a constant demand for alternative purification methodologies, to ameliorate the dependence on chromatography, while still adhering to regulatory concerns over product purity and safety. In this article, we present an up-to-date view of bioseparation, with emphasis on magnetic separation and its potential application in the field. Additionally, we discuss the economic and performance benefits of synthetic ligands, in the form of peptides and miniaturized antibody fragments, compared to full-length antibodies. We propose that adoption of synthetic affinity ligands coupled with magnetic adsorbents, will play an important role in enabling sustainable bioprocessing in the future. PMID:26032605

  5. 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.

  6. Dynamical surface affinity of diphasic liquids as a probe of wettability of multimodal porous media.

    PubMed

    Korb, J-P; Freiman, G; Nicot, B; Ligneul, P

    2009-12-01

    We introduce a method for estimating the wettability of rock/oil/brine systems using noninvasive in situ nuclear magnetic relaxation dispersion. This technique scans over a large range of applied magnetic fields and yields unique information about the extent to which a fluid is dynamically correlated with a solid rock surface. Unlike conventional transverse relaxation studies, this approach is a direct probe of the dynamical surface affinity of fluids. To quantify these features we introduce a microscopic dynamical surface affinity index which measures the dynamical correlation (i.e., the microscopic wettability) between the diffusive fluid and the fixed paramagnetic relaxation sources at the pore surfaces. We apply this method to carbonate reservoir rocks which are known to hold about two thirds of the world's oil reserves. Although this nondestructive method concerns here an application to rocks, it could be generalized as an in situ liquid/surface affinity indicator for any multimodal porous medium including porous biological media. PMID:20365175

  7. Affinity separation in magnetically stabilized fluidized beds: synthesis and performance of packing materials

    SciTech Connect

    Lochmueller, C.H.; Wigman, L.S.

    1987-11-01

    A magnetically stabilized fluidized-bed separator designed to test the use of pellicular, ferromagnetic affinity chromatography packing materials has been developed. A wire wound solenoid was used to produce the magnetic field. The ferromagnetic packing material is comprised of a magnetite-containing, polyurethane gel coated onto polystyrene beads. The gel contains free carboxyl groups. These were carbodiimide-coupled to soy trypsin inhibitor and the material used for trypsin purification. Narrow-band affinity chromatography was carried out in packed-bed, fluidized-bed, and magnetically stabilized, fluidized-bed separators. Pressure drop, capacity, dilution, and peak asymmetry were evaluated for each type of separator. The three types provide comparable efficiency but the fluidized separators exhibit a much lower pressure drop. As might be expected, fluidized-bed separators perform well for affinity chromatography (large k') but poorly for size exclusion chromatography.

  8. Wakimoto modules for the affine superalgebra sl( {overline2}/{1}) and noncritical N = 2 strings

    NASA Astrophysics Data System (ADS)

    Bowcock, P.; Koktava, R.-L. K.; Taormina, A.

    1996-02-01

    Free field representations of the affine superalgebra A(1, 0) (1) at level k corresponding to two inequivalent choices of the simple roots are shown to be related by nonlinear canonical field transformations, both at the classical and at the quantum level. The ambiguity in the choice of the Wakimoto module needed in the description of physical states in the noncritical N = 2 string is therefore lifted.

  9. Computational protocol for predicting the binding affinities of zinc containing metalloprotein-ligand complexes.

    PubMed

    Jain, Tarun; Jayaram, B

    2007-06-01

    Zinc is one of the most important metal ions found in proteins performing specific functions associated with life processes. Coordination geometry of the zinc ion in the active site of the metalloprotein-ligand complexes poses a challenge in determining ligand binding affinities accurately in structure-based drug design. We report here an all atom force field based computational protocol for estimating rapidly the binding affinities of zinc containing metalloprotein-ligand complexes, considering electrostatics, van der Waals, hydrophobicity, and loss in conformational entropy of protein side chains upon ligand binding along with a nonbonded approach to model the interactions of the zinc ion with all the other atoms of the complex. We examined the sensitivity of the binding affinity predictions to the choice of Lennard-Jones parameters, partial atomic charges, and dielectric treatments adopted for system preparation and scoring. The highest correlation obtained was R2 = 0.77 (r = 0.88) for the predicted binding affinity against the experiment on a heterogenous dataset of 90 zinc containing metalloprotein-ligand complexes consisting of five unique protein targets. Model validation and parameter analysis studies underscore the robustness and predictive ability of the scoring function. The high correlation obtained suggests the potential applicability of the methodology in designing novel ligands for zinc-metalloproteins. The scoring function has been web enabled for free access at www.scfbio-iitd.res.in/software/drugdesign/bapplz.jsp as BAPPL-Z server (Binding Affinity Prediction of Protein-Ligand complexes containing Zinc metal ions).

  10. 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.

  11. 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

  12. 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

  13. 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.

  14. 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.

  15. 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

  16. Negative homotropic cooperativity and affinity heterogeneity: preparation of yeast glyceraldehyde-3-phosphate dehydrogenase with maximal affinity homogeneity.

    PubMed Central

    Gennis, L S

    1976-01-01

    A three-step procedure including affinity chromatography on NAD+-azobenzamidopropyl-Sepharose has been designed for the purification of yeast glyceraldehyde-3-phosphate dehydrogenase [D-glyceraldehyde-3-phosphate: NAD+ oxidoreductase (phosphorylating), EC 1.2.1.12] with maximized specific activity and maximized homogeneity with respect to affinity for the coenzyme, NAD+.Binding isotherms allow the analysis of cooperativity patterns that disclose both the average ligand affinity in the system and the distribution of ligands among the sites, only for systems with complete affinity homogeneity. The presence of affinity heterogeneity, resulting from multiple oligomeric species differing only in their affinity for coenzyme, gives rise to isotherms which falsely manifest apparent negative cooperativity. A method for distinguishing negative homotropic cooperativity from affinity heterogeneity is suggested. PMID:186779

  17. 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

  18. Development of thermo-responsive hydrogels with immobilized metal affinity groups

    NASA Astrophysics Data System (ADS)

    Yoon, Young-Seo

    A Hydrogel is defined as a polymeric material which possesses the ability to swell in water and retain a significant fraction of water within its structure, but which will not dissolve in water. Hydrogels have been studied by many researchers because they have many useful applications in bio related fields such as drug delivery, bioseparation, and etc. In this thesis, a new hydrogel system that possesses the characteristics of thermo-responsive swelling property and immobilized metal affinity was developed. This affinity material consists of a hydrogel with stimuli responsive swelling characteristics to provide modulated diffusivity and size selectivity. Covalently bound ligands within hydrogels provide highly selective and tunable affinity-based separation. Swelling and affinity properties can be independently controlled by regulating the temperature or pH of the solution to provide a sequential separations scheme. The developed affinity hydrogels incorporate multiple modes of separations or recovery and concentrate specific solutes in chromatographic systems. Thermal sensitive affinity hydrogels were synthesized from a N-isopropylacrylamide (NIPAAm) monomer, a crosslinker (1,4-bismethylene acrylamide) and a ligand attachable co-monomer acrylamide (AAm), using free radical chemistry. The ligand of choice is the metal affinity iminodiacetic acid (IDA) which is bound to hydrogel backbone via a spacer arm. The challenge lay in incorporating affinity ligands without affecting the temperature induced swelling of the hydrogel. Thus, PNIPAAm-Am hydrogels are functionalized with a spacer arm (1,4-butanediol diglycidyl ether), the chelating ligand IDA and a divalent metal ion (Cu2+). This ligand binds histidine groups at high pH and releases them upon protonation of histidine at low pH. This can be used to separate proteins based on the occurrence of surface histidine residues in them. The resulting affinity hydrogel was shown to adsorb the protein chicken egg white

  19. Affinity chromatography with an immobilized RNA enzyme.

    PubMed Central

    Vioque, A; Altman, S

    1986-01-01

    M1 RNA, the catalytic subunit of Escherichia coli RNase P, has been covalently linked at its 3' terminus to agarose beads. Unlike M1 RNA, which is active in solution in the absence of the protein component (C5) of RNase P, the RNA linked to the beads is active only in the presence of C5 protein. Affinity chromatography of crude extracts of E. coli on a column prepared from the beads to which the RNA has been crosslinked results in the purification of C5 protein in a single step. The protein has been purified in this manner from cells that contain a plasmid, pINIIIR20, which includes the gene that codes for C5 protein. A 6-fold amplification of the expression of C5 protein is found in these cells after induction as compared to cells that do not harbor the plasmid. Images PMID:3526344

  20. 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.

  1. High-affinity neuropeptide Y receptor antagonists.

    PubMed Central

    Daniels, A J; Matthews, J E; Slepetis, R J; Jansen, M; Viveros, O H; Tadepalli, A; Harrington, W; Heyer, D; Landavazo, A; Leban, J J

    1995-01-01

    Neuropeptide Y (NPY) is one of the most abundant peptide transmitters in the mammalian brain. In the periphery it is costored and coreleased with norepinephrine from sympathetic nerve terminals. However, the physiological functions of this peptide remain unclear because of the absence of specific high-affinity receptor antagonists. Three potent NPY receptor antagonists were synthesized and tested for their biological activity in in vitro, ex vivo, and in vivo functional assays. We describe here the effects of these antagonists inhibiting specific radiolabeled NPY binding at Y1 and Y2 receptors and antagonizing the effects of NPY in human erythroleukemia cell intracellular calcium mobilization perfusion pressure in the isolated rat kidney, and mean arterial blood pressure in anesthetized rats. PMID:7568074

  2. Evaluation system of negative electron affinity photocathode

    NASA Astrophysics Data System (ADS)

    Fu, Rongguo; Chang, Benkang; Qian, Yunsheng; Wang, Guihua; Zong, Zhiyuan

    2001-10-01

    This article first describes the background of the research and manufacture of evaluation system of Negative Electron Affinity photocathode. This article designs a set of super high vacuum system for activating NEA photocathode on the base of activation theory, the process of design and debugging is given. The system is composed of three parts: super high vacuum system for GaAs material activation, multi-meter testing system, surface analysis system. The system is used for on-line evaluation of activating of NEA photocathode. The technical parameters and structure of the evaluation system of NEA photocathode are given in the paper. The system is finished and experiments are made. At last the picture of the system is given.

  3. 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

  4. Purification of glycolytic enzymes by using affinity-elution chromatography.

    PubMed Central

    Scopes, R K

    1977-01-01

    1. A systematic procedure for the purification of enzymes by affinity-elution chromatography is described. Enzymes are adsorbed on a cation-exchanger, and eluted with ligands specific for the enzyme concerned. 2. All of the glycolytic and some related enzymes present in rabbit muscle can be purified by the affinity-elution technique. The pH range for adsorption and elution of each enzyme was found, and the effects of minor variations of conditions are described. 3. A description of experimental conditions suitable for affinity elution of each enzyme is given, together with special features relevant to each individual enzyme. 4. Theoretical considerations of affinity elution chromatography are discussed, including its limitations, advantages and disadvantages compared with affinity-adsorption chromatography. Possible developments are suggested to cover enzymes which because of their adsorption characteristics are not at present amenable to affinity-elution procedures. PMID:192194

  5. Antibody-based affinity cryo-EM grid.

    PubMed

    Yu, Guimei; Li, Kunpeng; Jiang, Wen

    2016-05-01

    The Affinity Grid technique combines sample purification and cryo-Electron Microscopy (cryo-EM) grid preparation into a single step. Several types of affinity surfaces, including functionalized lipids monolayers, streptavidin 2D crystals, and covalently functionalized carbon surfaces have been reported. More recently, we presented a new affinity cryo-EM approach, cryo-SPIEM, which applies the traditional Solid Phase Immune Electron Microscopy (SPIEM) technique to cryo-EM. This approach significantly simplifies the preparation of affinity grids and directly works with native macromolecular complexes without need of target modifications. With wide availability of high affinity and high specificity antibodies, the antibody-based affinity grid would enable cryo-EM studies of the native samples directly from cell cultures, targets of low abundance, and unstable or short-lived intermediate states.

  6. 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.

  7. Affinity chromatography for purification of two urokinases from human urine.

    PubMed

    Takahashi, R; Akiba, K; Koike, M; Noguchi, T; Ezure, Y

    2000-05-26

    A new affinity chromatography (hydrophobic-mediated affinity chromatography), which was characterized by the matrix having both affinity site to urokinase and hydrophobic site, was established for the purification of urokinase from human urine. The hydrophobic affinity matrix (tentatively named PAS in the text) was prepared by immobilizing 6-aminocaproic acid on Sepharose CL-6B, followed by a coupling p-aminobenzamidine to a part of the hydrophobic site on the matrix. The PAS matrix was applied to the purification of urokinase from human urine, and high- and low-molecular weight pure urokinases were efficiently obtained in high yield by the present method. PMID:10892585

  8. 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

  9. Multiplexed protein profiling by sequential affinity capture

    PubMed Central

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

    2016-01-01

    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

  10. 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.

  11. Prostate Cancer and Bone: The Elective Affinities

    PubMed Central

    2014-01-01

    The onset of metastases dramatically changes the prognosis of prostate cancer patients, determining increased morbidity and a drastic fall in survival expectancy. Bone is a common site of metastases in few types of cancer, and it represents the most frequent metastatic site in prostate cancer. Of note, the prevalence of tumor relapse to the bone appears to be increasing over the years, likely due to a longer overall survival of prostate cancer patients. Bone tropism represents an intriguing challenge for researchers also because the preference of prostate cancer cells for the bone is the result of a sequential series of targetable molecular events. Many factors have been associated with the peculiar ability of prostate cancer cells to migrate in bone marrow and to determine mixed osteoblastic/osteolytic lesions. As anticipated by the success of current targeted therapy aimed to block bone resorption, a better understanding of molecular affinity between prostate cancer and bone microenvironment will permit us to cure bone metastasis and to improve prognosis of prostate cancer patients. PMID:24971315

  12. 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\

  13. Current Understanding of the Binding Sites, Capacity, Affinity, and Biological Significance of Metals in Melanin

    PubMed Central

    Hong, Lian; Simon, John D.

    2008-01-01

    Metal chelation is often invoked as one of the main biological functions of melanin. In order to understand the interaction between metals and melanin, extensive studies have been carried out to determine the nature of the metal binding sites, binding capacity and affinity. These data are central to efforts aimed at elucidating the role metal binding plays in determining the physical, structural, biological, and photochemical properties of melanin. This article examines the current state of understanding of this field. PMID:17580858

  14. 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…

  15. 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.

  16. 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'…

  17. High affinity retinoic acid receptor antagonists: analogs of AGN 193109.

    PubMed

    Johnson, A T; Wang, L; Gillett, S J; Chandraratna, R A

    1999-02-22

    A series of high affinity retinoic acid receptor (RAR) antagonists were prepared based upon the known antagonist AGN 193109 (2). Introduction of various phenyl groups revealed a preference for substitution at the para-position relative to the meta-site. Antagonists with the highest affinities for the RARs possessed hydrophobic groups, however, the presence of polar functionality was also well tolerated.

  18. Affinity Regulates Spatial Range of EGF Receptor Autocrine Ligand Binding

    SciTech Connect

    Dewitt, Ann; Iida, Tomoko; Lam, Ho-Yan; Hill, Virginia; Wiley, H S.; Lauffenburger, Douglas A.

    2002-08-08

    Proper spatial localization of EGFR signaling activated by autocrine ligands represents a critical factor in embryonic development as well as tissue organization and function, and ligand/receptor binding affinity is among the molecular and cellular properties suggested to play a role in governing this localization. The authors employ a computational model to predict how receptor-binding affinity affects local capture of autocrine ligand vis-a-vis escape to distal regions, and provide experimental test by constructing cell lines expressing EGFR along with either wild-type EGF or a low-affinity mutant, EGF{sup L47M}. The model predicts local capture of a lower affinity autocrine ligand to be less efficient when the ligand production rate is small relative to receptor appearance rate. The experimental data confirm this prediction, demonstrating that cells can use ligand/receptor binding affinity to regulate ligand spatial distribution when autocrine ligand production is limiting for receptor signaling.

  19. Detection of protein-protein interactions using tandem affinity purification.

    PubMed

    Goodfellow, Ian; Bailey, Dalan

    2014-01-01

    Tandem affinity purification (TAP) is an invaluable technique for identifying interaction partners for an affinity tagged bait protein. The approach relies on the fusion of dual tags to the bait before separate rounds of affinity purification and precipitation. Frequently two specific elution steps are also performed to increase the specificity of the overall technique. In the method detailed here, the two tags used are protein G and a short streptavidin binding peptide; however, many variations can be employed. In our example the tags are separated by a cleavable tobacco etch virus protease target sequence, allowing for specific elution after the first round of affinity purification. Proteins isolated after the final elution step in this process are concentrated before being identified by mass spectrometry. The use of dual affinity tags and specific elution in this technique dramatically increases both the specificity and stringency of the pull-downs, ensuring a low level of background nonspecific interactions.

  20. 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.

  1. Affinity based information diffusion model in social networks

    NASA Astrophysics Data System (ADS)

    Liu, Hongli; Xie, Yun; Hu, Haibo; Chen, Zhigao

    2014-12-01

    There is a widespread intuitive sense that people prefer participating in spreading the information in which they are interested. The affinity of people with information disseminated can affect the information propagation in social networks. In this paper, we propose an information diffusion model incorporating the mechanism of affinity of people with information which considers the fitness of affinity values of people with affinity threshold of the information. We find that the final size of information diffusion is affected by affinity threshold of the information, average degree of the network and the probability of people's losing their interest in the information. We also explore the effects of other factors on information spreading by numerical simulations and find that the probabilities of people's questioning and confirming the information can affect the propagation speed, but not the final scope.

  2. 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

  3. 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

  4. Enzyme-gold affinity labelling of cellulose.

    PubMed

    Berg, R H; Erdos, G W; Gritzali, M; Brown, R D

    1988-04-01

    The enzyme-linked colloidal gold affinity labelling technique was tested as a method to localize cellulose on thin sections of plant cell walls and slime mold spores. Commercially available cellulase from cultures of Trichoderma reesei, the main components being cellobiohydrolase I and II (CBH I, CBH II) and endoglucanase (EG), was linked to colloidal gold by using standard techniques and applied as a dilute, buffered suspension to thin sections. After brief exposure, e.g., 15-30 minutes, cellulose exposed on the surface of sections was labelled with the enzyme-gold complex. Poststaining did not appear to have a deleterious effect on the labelled sections. The specificity of labelling was demonstrated by its complete inhibition when carboxymethylcellulose was incorporated in the labelling mixture, by lack of labelling of 1,4-beta-mannans or 1,3-beta-xylans in noncellulosic walls of marine algae, by lack of labelling of 1,4-beta-glucans in chitin, by much lower labelling density when done at 4 degrees C, and by lack of labelling when sections were predigested with cellulase. Labelling with the crude commercial cellulase was compared to labelling with purified CBH I-, CBH II-, and EG-linked colloidal gold, and the labelling pattern was similar. This method was found useful on conventionally fixed material and required no special preparation other than the use of inert (Ni or Au) grids and 0.5% gelatin to reduce nonspecific binding of the gold complex. Labelling was similar in the several embedding resins tested: LR White, Lowicryl K4M, Epon 812, and Spurr's.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. Optimal affine-invariant matching: performance characterization

    NASA Astrophysics Data System (ADS)

    Costa, Mauro S.; Haralick, Robert M.; Shapiro, Linda G.

    1992-04-01

    The geometric hashing scheme proposed by Lamdan and Wolfson can be very efficient in a model-based matching system, not only in terms of the computational complexity involved, but also in terms of the simplicity of the method. In a recent paper, we discussed errors that can occur with this method due to quantization, stability, symmetry, and noise problems. These errors make the original geometric hashing technique unsuitable for use on the factory floor. Beginning with an explicit noise model, which the original Lamdan and Wolfson technique lacks, we derived an optimal approach that overcomes these problems. We showed that the results obtained with the new algorithm are clearly better than the results from the original method. This paper addresses the performance characterization of the geometric hashing technique, more specifically the affine-invariant point matching, applied to the problem of recognizing and determining the pose of sheet metal parts. The experiments indicate that with a model having 10 to 14 points, with 2 points of the model undetected and 10 extraneous points detected, and with the model points perturbed by Gaussian noise of standard deviation 3 (0.58 of range), the average amount of computation required to obtain an answer is equivalent to trying 11 of the possible three-point bases. The misdetection rate, measured by the percentage of correct bases matches that fail to verify, is 0.9. The percentage of incorrect bases that successfully produced a match that did verify (false alarm rate) is 13. And, finally, 2 of the experiments failed to find a correct match and verify it. Results for experiments with real images are also presented.

  6. 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

  7. Surface-modified magnetic colloids for affinity adsorption of immunoglobulins

    NASA Astrophysics Data System (ADS)

    Martins, Fernanda; Pinho, Samantha C.; Zollner, Terezinha C. A.; Zollner, Ricardo L.; de Cuyper, Marcel; Santana, Maria Helena A.

    This work describes the preparation, characterization and in vitro adsorption tests of surface-modified magnetoliposomes for affinity binding of (i) anticardiolipin (isotype G) antibodies and (ii) specific isotype E antibodies generated by hypersensitivity reactions in humans with respiratory allergy. In the first case, cardiolipin embedded in the bilayer of magnetoliposomes was used as specific ligand. In the second case, antigenic proteins present in an extract of Dermatophagoids pteronyssinus and Blomia tropicalis mites were covalently coupled on the surface of magnetoliposomes via a diglycolic spacer arm, and used as specific ligands for IgE. Antibody adsorption was performed in a high-gradient magnetophoresis system, using either sera of healthy individuals or a pool of sera from autoimmune or allergic patients. The selectivity and capacity of the system were quantified by a frontal analysis in a capillary column, and by constructing breakthrough curves. The results show that the highest yield and selectivity were obtained if the ligand was extended into the aqueous layer surrounding the magnetoliposome surface. A 100% selectivity was obtained for adsorption of specific IgE, and 8% for IgG. These results demonstrate the potentialities of both types of surface-modified magnetic biocolloids in the field of in vitro diagnosis tests for allergic or autoimmune conditions.

  8. How Structure Defines Affinity in Protein-Protein Interactions

    PubMed Central

    Erijman, Ariel; Rosenthal, Eran; Shifman, Julia M.

    2014-01-01

    Protein-protein interactions (PPI) in nature are conveyed by a multitude of binding modes involving various surfaces, secondary structure elements and intermolecular interactions. This diversity results in PPI binding affinities that span more than nine orders of magnitude. Several early studies attempted to correlate PPI binding affinities to various structure-derived features with limited success. The growing number of high-resolution structures, the appearance of more precise methods for measuring binding affinities and the development of new computational algorithms enable more thorough investigations in this direction. Here, we use a large dataset of PPI structures with the documented binding affinities to calculate a number of structure-based features that could potentially define binding energetics. We explore how well each calculated biophysical feature alone correlates with binding affinity and determine the features that could be used to distinguish between high-, medium- and low- affinity PPIs. Furthermore, we test how various combinations of features could be applied to predict binding affinity and observe a slow improvement in correlation as more features are incorporated into the equation. In addition, we observe a considerable improvement in predictions if we exclude from our analysis low-resolution and NMR structures, revealing the importance of capturing exact intermolecular interactions in our calculations. Our analysis should facilitate prediction of new interactions on the genome scale, better characterization of signaling networks and design of novel binding partners for various target proteins. PMID:25329579

  9. The sodium ion affinities of asparagine, glutamine, histidine and arginine

    NASA Astrophysics Data System (ADS)

    Wang, Ping; Ohanessian, Gilles; Wesdemiotis, Chrys

    2008-01-01

    The sodium ion affinities of the amino acids Asn, Gln, His and Arg have been determined by experimental and computational approaches (for Asn, His and Arg). Na+-bound heterodimers with amino acid and peptide ligands (Pep1, Pep2) were produced by electrospray ionization. From the dissociation kinetics of these Pep1-Na+-Pep2 ions to Pep1-Na+ and Pep2-Na+, determined by collisionally activated dissociation, a ladder of relative affinities was constructed and subsequently converted to absolute affinities by anchoring the relative values to known Na+ affinities. The Na+ affinities of Asn, His and Arg, were calculated at the MP2(full)/6-311+G(2d,2p)//MP2/6-31G(d) level of ab initio theory. The resulting experimental and computed Na+ affinities are in excellent agreement with one another. These results, combined with those of our previous studies, yield the sodium ion affinities of 18 out of the 20 [alpha]-amino acids naturally occurring in peptides and proteins of living systems.

  10. 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

  11. Affine differential geometry analysis of human arm movements.

    PubMed

    Flash, Tamar; Handzel, Amir A

    2007-06-01

    Humans interact with their environment through sensory information and motor actions. These interactions may be understood via the underlying geometry of both perception and action. While the motor space is typically considered by default to be Euclidean, persistent behavioral observations point to a different underlying geometric structure. These observed regularities include the "two-thirds power law", which connects path curvature with velocity, and "local isochrony", which prescribes the relation between movement time and its extent. Starting with these empirical observations, we have developed a mathematical framework based on differential geometry, Lie group theory and Cartan's moving frame method for the analysis of human hand trajectories. We also use this method to identify possible motion primitives, i.e., elementary building blocks from which more complicated movements are constructed. We show that a natural geometric description of continuous repetitive hand trajectories is not Euclidean but equi-affine. Specifically, equi-affine velocity is piecewise constant along movement segments, and movement execution time for a given segment is proportional to its equi-affine arc-length. Using this mathematical framework, we then analyze experimentally recorded drawing movements. To examine movement segmentation and classification, the two fundamental equi-affine differential invariants-equi-affine arc-length and curvature are calculated for the recorded movements. We also discuss the possible role of conic sections, i.e., curves with constant equi-affine curvature, as motor primitives and focus in more detail on parabolas, the equi-affine geodesics. Finally, we explore possible schemes for the internal neural coding of motor commands by showing that the equi-affine framework is compatible with the common model of population coding of the hand velocity vector when combined with a simple assumption on its dynamics. We then discuss several alternative explanations

  12. 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.

  13. 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.

  14. Complementary metal oxide semiconductor-compatible silicon nanowire biofield-effect transistors as affinity biosensors.

    PubMed

    Duan, Xuexin; Rajan, Nitin K; Izadi, Mohammad Hadi; Reed, Mark A

    2013-11-01

    Affinity biosensors use biorecognition elements and transducers to convert a biochemical event into a recordable signal. They provides the molecule binding information, which includes the dynamics of biomolecular association and dissociation, and the equilibrium association constant. Complementary metal oxide semiconductor-compatible silicon (Si) nanowires configured as a field-effect transistor (NW FET) have shown significant advantages for real-time, label-free and highly sensitive detection of a wide range of biomolecules. Most research has focused on reducing the detection limit of Si-NW FETs but has provided less information about the real binding parameters of the biomolecular interactions. Recently, Si-NW FETs have been demonstrated as affinity biosensors to quantify biomolecular binding affinities and kinetics. They open new applications for NW FETs in the nanomedicine field and will bring such sensor technology a step closer to commercial point-of-care applications. This article summarizes the recent advances in bioaffinity measurement using Si-NW FETs, with an emphasis on the different approaches used to address the issues of sensor calibration, regeneration, binding kinetic measurements, limit of detection, sensor surface modification, biomolecule charge screening, reference electrode integration and nonspecific molecular binding.

  15. Infinitesimal affine automorphisms of symplectic connections

    NASA Astrophysics Data System (ADS)

    Fox, Daniel J. F.

    2016-08-01

    Conditions are given under which an infinitesimal automorphism of a torsion-free connection preserving a symplectic form is necessarily a symplectic vector field. An example is given of a compact symplectic nilmanifold admitting a flat symplectic connection and an infinitesimal automorphism that is not symplectic.

  16. Real-time affine invariant gesture recognition for LED smart lighting control

    NASA Astrophysics Data System (ADS)

    Chen, Xu; Liao, Miao; Feng, Xiao-Fan

    2015-03-01

    Gesture recognition has attracted extensive research interest in the field of human computer interaction. Realtime affine invariant gesture recognition is an important and challenging problem. This paper presents a robust affine view invariant gesture recognition system for realtime LED smart light control. As far as we know, this is the first time that gesture recognition has been applied for control LED smart light in realtime. Employing skin detection, hand blobs captured from a top view camera are first localized and aligned. Subsequently, SVM classifiers trained on HOG features and robust shape features are then utilized for gesture recognition. By accurately recognizing two types of gestures ("gesture 8" and a "5 finger gesture"), a user is enabled to toggle lighting on/off efficiently and control light intensity on a continuous scale. In each case, gesture recognition is rotation- and translation-invariant. Extensive evaluations in an office setting demonstrate the effectiveness and robustness of the proposed gesture recognition algorithm.

  17. Dense Stereo Matching Method Based on Local Affine Model.

    PubMed

    Li, Jie; Shi, Wenxuan; Deng, Dexiang; Jia, Wenyan; Sun, Mingui

    2013-07-01

    A new method for constructing an accurate disparity space image and performing an efficient cost aggregation in stereo matching based on local affine model is proposed in this paper. The key algorithm includes a new self-adapting dissimilarity measurement used for calculating the matching cost and a local affine model used in cost aggregation stage. Different from the traditional region-based methods, which try to change the matching window size or to calculate an adaptive weight to do the aggregation, the proposed method focuses on obtaining the efficient and accurate local affine model to aggregate the cost volume while preserving the disparity discontinuity. Moreover, the local affine model can be extended to the color space. Experimental results demonstrate that the proposed method is able to provide subpixel precision disparity maps compared with some state-of-the-art stereo matching methods. PMID:24163727

  18. On the thermodynamic basis of the affinity decay rate

    NASA Astrophysics Data System (ADS)

    Garcia-Colín, L. S.; Piña, E.; de la Selva, S. M. T.

    1990-03-01

    In the past five years exhaustive studies in chemical reactions have lead to an empirical equation describing how isothermal-isometric homogeneous reactions evolve towards equilibrium independently of their particular mechanism or rate law. Such an equation expresses the time rate of change of the chemical affinity as a linear function of the inverse of time. In this paper we show that by invoking the local equilibrium hypothesis one may provide, a time evolution equation for the chemical affinity that is uniquely given by the solution of the particular rate law of the reaction considered. Consequently such an equation is not of the same functional form for all reactions. On the other hand, integration of Dalton's law under specific initial conditions, together with the local equilibrium assumption and the ideality requirement for the reacting species, exhibits a unique inverse time decay for the chemical affinity. This explains the good fitting of the inverse in time dependence of the chemical affinity with experimental data.

  19. A thermodynamic approach to the affinity optimization of drug candidates.

    PubMed

    Freire, Ernesto

    2009-11-01

    High throughput screening and other techniques commonly used to identify lead candidates for drug development usually yield compounds with binding affinities to their intended targets in the mid-micromolar range. The affinity of these molecules needs to be improved by several orders of magnitude before they become viable drug candidates. Traditionally, this task has been accomplished by establishing structure activity relationships to guide chemical modifications and improve the binding affinity of the compounds. As the binding affinity is a function of two quantities, the binding enthalpy and the binding entropy, it is evident that a more efficient optimization would be accomplished if both quantities were considered and improved simultaneously. Here, an optimization algorithm based upon enthalpic and entropic information generated by Isothermal Titration Calorimetry is presented.

  20. 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

  1. 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

  2. Hydride affinities of cumulated, isolated, and conjugated dienes in acetonitrile.

    PubMed

    Zhu, Xiao-Qing; Liang, Hao; Zhu, Yan; Cheng, Jin-Pei

    2008-11-01

    The hydride affinities (defined as the enthalpy changes in this work) of 15 polarized dienes [five phenyl sulfone substituted allenes (1a), the corresponding five isolated dienes (1b), and the corresponding five conjugated dienes (1c)] in acetonitrile solution were determined by titration calorimetry for the first time. The results display that the hydride affinity scales of the 15 dienes in acetonitrile range from -71.6 to -73.9 kcal/mol for 1a, from -46.2 to -49.7 kcal/mol for 1b, and from -45.0 to -46.5 kcal/mol for 1c, which indicates that the hydride-obtaining abilities of the cumulated dienes (1a) are not only much larger than those of the corresponding conjugated dienes (1c) but also much larger than those of the corresponding isolated dienes (1b). The hydrogen affinities of the 15 dienes as well as the hydrogen affinities and the proton affinities of the radical anions of the dienes (1(-*)) in acetonitrile were also evaluated by using relative thermodynamic cycles according to Hess's law. The results show that (i) the hydrogen affinities of the neutral dienes 1 cover a range from -44.5 to -45.6 kcal/mol for 1a, from -20.4 to -21.4 kcal/mol for 1b, and from -17.3 to -18.5 kcal/mol for 1c; (ii) the hydrogen affinities of the radical anions of the dienes (1(-*)) in acetonitrile cover a range from -40.6 to -47.2 kcal/mol for 1a(-*), from -21.6 to -29.6 kcal/mol for 1b(-*), and from -10.0 to -15.4 kcal/mol for 1c(-*); (iii) the proton affinities of the 15 1a(-*) in acetonitrile cover a range from -97.0 to -100.6 kcal/mol for 1a(-*), from -77.8 to -83.4 kcal/mol for 1b(-*), and from -66.2 to -68.9 kcal/mol for 1c(-*). The main reasons for the great difference between the cumulated dienes and the corresponding isolated and conjugated dienes in the hydride affinity, hydrogen affinity, and proton affinity have been examined. It is evident that these experimental results should be quite valuable to facilitate the elucidation of the origins of the especially high

  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. 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

  5. Affinity Density: a novel genomic approach to the identification of transcription factor regulatory targets

    PubMed Central

    Hazelett, Dennis J.; Lakeland, Daniel L.; Weiss, Joseph B.

    2009-01-01

    Methods: A new method was developed for identifying novel transcription factor regulatory targets based on calculating Local Affinity Density. Techniques from the signal-processing field were used, in particular the Hann digital filter, to calculate the relative binding affinity of different regions based on previously published in vitro binding data. To illustrate this approach, the complete genomes of Drosophila melanogaster and D.pseudoobscura were analyzed for binding sites of the homeodomain proteinc Tinman, an essential heart development gene in both Drosophila and Mouse. The significant binding regions were identified relative to genomic background and assigned to putative target genes. Valid candidates common to both species of Drosophila were selected as a test of conservation. Results: The new method was more sensitive than cluster searches for conserved binding motifs with respect to positive identification of known Tinman targets. Our Local Affinity Density method also identified a significantly greater proportion of Tinman-coexpressed genes than equivalent, optimized cluster searching. In addition, this new method predicted a significantly greater than expected number of genes with previously published RNAi phenotypes in the heart. Availability: Algorithms were implemented in Python, LISP, R and maxima, using MySQL to access locally mirrored sequence data from Ensembl (D.melanogaster release 4.3) and flybase (D.pseudoobscura). All code is licensed under GPL and freely available at http://www.ohsu.edu/cellbio/dev_biol_prog/affinitydensity/. Contact: hazelett@ohsu.edu PMID:19401399

  6. 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

  7. Affinity monolith chromatography: A review of principles and recent analytical applications

    PubMed Central

    Pfaunmiller, Erika L.; Paulemond, Marie Laura; Dupper, Courtney M.; Hage, David S.

    2012-01-01

    Affinity monolith chromatography (AMC) is a type of liquid chromatography that uses a monolithic support and a biologically-related binding agent as a stationary phase. AMC is a powerful method for the selective separation, analysis or studies of specific target compounds in a sample. This review discusses the basic principles of AMC and recent developments or applications of this method, with particular emphasis being given to work that has appeared in the last five years. Various materials that have been used to prepare columns for AMC are examined, including organic monoliths, silica monoliths, agarose monoliths and cryogels. These supports have been used in AMC for formats that have ranged from traditional columns to disks, microcolumns and capillaries. Many binding agents have also been employed in AMC, such as antibodies, enzymes, proteins, lectins, immobilized metal-ions and dyes. Some applications that have been reported with these binding agents in AMC are bioaffinity chromatography, immunoaffinity chromatography or immunoextraction, immobilized metal-ion affinity chromatography, dye-ligand affinity chromatography, chiral separations and biointeraction studies. Examples are presented from fields that include analytical chemistry, pharmaceutical analysis, clinical testing and biotechnology. Current trends and possible future directions in AMC are also discussed. PMID:23187827

  8. 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

  9. Gradient-based habitat affinities predict species vulnerability to drought.

    PubMed

    Debinski, Diane M; Caruthers, Jennet C; Cook, Dianne; Crowley, Jason; Wickham, Hadley

    2013-05-01

    Ecological fingerprints of climate change are becoming increasingly evident at broad geographical scales as measured by species range shifts and changes in phenology. However, finer-scale species-level responses to environmental fluctuations may also provide an important bellwether of impending future community responses. Here we examined changes in abundance of butterfly species along a hydrological gradient of six montane meadow habitat types in response to drought. Our data collection began prior to the drought, and we were able to track changes for 11 years, of which eight were considered mild to extreme drought conditions. We separated the species into those that had an affinity for hydric vs. xeric habitats. We suspected that drought would favor species with xeric habitat affinities, but that there could be variations in species-level responses along the hydrological gradient. We also suspected that mesic meadows would be most sensitive to drought conditions. Temporal trajectories were modeled for both species groups (hydric vs. xeric affinity) and individual species. Abundances of species with affinity for xeric habitats increased in virtually all meadow types. Conversely, abundances of species with affinity for hydric habitats decreased, particularly in mesic and xeric meadows. Mesic meadows showed the most striking temporal abundance trajectory: Increasing abundances of species with xeric habitat affinity were offset by decreasing or stable abundances of species with hydric habitat affinity. The one counterintuitive finding was that, in some hydric meadows, species with affinity for hydric habitats increased. In these cases, we suspect that decreasing moisture conditions in hydric meadows actually increased habitat suitability because sites near the limit of moisture extremes for some species became more acceptable. Thus, species responses were relatively predictable based upon habitat affinity and habitat location along the hydrological gradient, and

  10. Effectors of hemoglobin. Separation of allosteric and affinity factors.

    PubMed Central

    Marden, M C; Bohn, B; Kister, J; Poyart, C

    1990-01-01

    The relative contributions of the allosteric and affinity factors toward the change in p50 have been calculated for a series of effectors of hemoglobin (Hb). Shifts in the ligand affinity of deoxy Hb and the values for 50% ligand saturation (p50) were obtained from oxygen equilibrium data. Because the high-affinity parameters (liganded conformation) are poorly determined from the equilibrium curves, they were determined from kinetic measurements of the association and dissociation rates with CO as ligand. The CO on-rates were obtained by flash photolysis measurements. The off-rates were determined from the rate of oxidation of HbCO by ferricyanide, or by replacement of CO with NO. The partition function of fully liganded hemoglobin for oxygen and CO is only slightly changed by the effectors. Measurements were made in the presence of the effectors 2,3-diphosphoglycerate (DPG), inositol hexakisphosphate (IHP), bezafibrate (Bzf), and two recently synthesized derivatives of Bzf (LR16 and L35). Values of p50 change by over a factor of 60; the on-rates decrease by nearly a factor of 8, with little change in the off-rates for the liganded conformation. The data indicate that both allosteric and affinity parameters are changed by the effectors; the changes in ligand affinity represent the larger contribution toward shifts in p50. PMID:2306490

  11. Melanin affinity and its possible role in neurodegeneration.

    PubMed

    Karlsson, Oskar; Lindquist, Nils Gunnar

    2013-12-01

    Certain drugs with melanin affinity are known to have caused pigmentary lesions in the eye and skin. This was the basis for the hypothesis that compounds with melanin affinity may cause damage also in other melanin-bearing tissues such as the substantia nigra. The heterogeneity of compounds that binds to melanin is large. Toxins, drugs, and several other compounds have melanin affinity. Compounds showing the highest affinity are mainly organic amines and metal ions. The binding of toxicants to melanin probably protects the cells initially. However, the binding is normally, slowly reversible and melanin may accumulate the toxicant and gradually release it into the cytosol. Several studies indicate that neuromelanin may play a significant role both in the initiation and in the progression of neurodegeneration. MPTP/MPP(+) that has been causally linked with Parkinsonism has high affinity for neuromelanin, and the induced dopaminergic denervation correlates with the neuromelanin content in the cells. This shows that the toxicological implications of the accumulation of toxicants in pigmented neurons and its possible role in neurodegeneration should not be neglected. Extracellular neuromelanin has been reported to activate dendritic cells and microglia. An initial neuronal damage induced by a neurotoxicant that leaks neuromelanin from the cells may therefore lead to a vicious cycle of neuroinflammation and further neurodegeneration. Although there are many clues to the particular vulnerability of dopaminergic neurons of substantia nigra in Parkinson's disease, the critical factors are not known. Further studies to determine the importance of neuromelanin in neurodegeneration and Parkinson's disease are warranted.

  12. 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

  13. Improving antibody binding affinity and specificity for therapeutic development.

    PubMed

    Bostrom, Jenny; Lee, Chingwei V; Haber, Lauric; Fuh, Germaine

    2009-01-01

    Affinity maturation is an important part of the therapeutic antibody development process as in vivo activity often requires high binding affinity. Here, we describe a targeted approach for affinity improvement of therapeutic antibodies. Sets of CDR residues that are solvent accessible and relatively diverse in natural antibodies are targeted for diversification. Degenerate oligonucleotides are used to generate combinatorial phage-displayed antibody libraries with varying degree of diversity at randomized positions from which high-affinity antibodies can be selected. An advantage of using antibodies for therapy is their exquisite target specificity, which enables selective antigen binding and reduces off-target effects. However, it can be useful, and often it is necessary, to generate cross-reactive antibodies binding to not only the human antigen but also the corresponding non-human primate or rodent orthologs. Such cross-reactive antibodies can be used to validate the therapeutic targeting and examine the safety profile in preclinical animal models before committing to a costly development track. We show how affinity improvement and cross-species binding can be achieved in a one-step process.

  14. Crystal Structure of A Plant Dual-Affinity Nitrate Transporter

    PubMed Central

    Sun, Ji; Bankston, John R.; Payandeh, Jian; Hinds, Thomas R.; Zagotta, William N.; Zheng, Ning

    2014-01-01

    Nitrate is a primary nutrient for plant growth, but its levels in soil can fluctuate by several orders of magnitude. Previous studies have identified Arabidopsis NRT1.1 as a dual-affinity nitrate transporter, which can take up nitrate over a wide range of concentrations. The mode of action of NRT1.1 is controlled by phosphorylation of a key residue, Thr101. Yet how this posttranslational modification switches the transporter between two affinity states remains unclear. Here we report the crystal structure of unphosphorylated NRT1.1, which reveals an unexpected homodimer in the inward-facing conformation. In this low-affinity state, the Thr101 phosphorylation site is embedded in a pocket immediately adjacent to the dimer interface, linking the phosphorylation status of the transporter to its oligomeric state. Using a cell-based fluorescence resonance energy transfer assay, we show that functional NRT1.1 indeed dimerizes in the cell membrane and the phosphomimetic mutation of Thr101 converts the protein into a monophasic high affinity transporter by structurally decoupling the dimer. Together with analyses of the substrate transport tunnel, our results establish a phosphorylation-controlled dimerization switch that allows NRT1.1 to uptake nitrate with two distinct affinity modes. PMID:24572362

  15. Fluorogen-Activating-Proteins as Universal Affinity Biosensors for Immunodetection

    PubMed Central

    Gallo, Eugenio; Vasilev, Kalin V.; Jarvik, Jonathan

    2014-01-01

    Fluorogen-activating-proteins (FAPs) are a novel platform of fluorescence biosensors utilized for protein discovery. The technology currently demands molecular manipulation methods that limit its application and adaptability. Here, we highlight an alternative approach based on universal affinity reagents for protein detection. The affinity reagents were engineered as bi-partite fusion proteins, where the specificity moiety is derived from IgG-binding proteins –Protein-A or Protein-G – and the signaling element is a FAP. In this manner, primary antibodies provide the antigenic selectivity against a desired protein in biological samples, while FAP affinity reagents target the constant region (Fc) of antibodies and provide the biosensor component of detection. Fluorescence results using various techniques indicate minimal background and high target specificity for exogenous and endogenous proteins in mammalian cells. Additionally, FAP-based affinity reagents provide enhanced properties of detection previously absent using conventional affinity systems. Distinct features explored in this report include: (1) unfixed signal wavelengths (excitation and emission) determined by the particular fluorogen chosen, (2) real-time user controlled fluorescence on-set and off-set, (3) signal wavelength substitution while performing live analysis, and (4) enhanced resistance to photobleaching. PMID:24122476

  16. Affinity Purification of Sequence-Specific DNA Binding Proteins

    NASA Astrophysics Data System (ADS)

    Kadonaga, James T.; Tjian, Robert

    1986-08-01

    We describe a method for affinity purification of sequence-specific DNA binding proteins that is fast and effective. Complementary chemically synthesized oligodeoxynucleotides that contain a recognition site for a sequence-specific DNA binding protein are annealed and ligated to give oligomers. This DNA is then covalently coupled to Sepharose CL-2B with cyanogen bromide to yield the affinity resin. A partially purified protein fraction is combined with competitor DNA and subsequently passed through the DNA-Sepharose resin. The desired sequence-specific DNA binding protein is purified because it preferentially binds to the recognition sites in the affinity resin rather than to the nonspecific competitor DNA in solution. For example, a protein fraction that is enriched for transcription factor Sp1 can be further purified 500- to 1000-fold by two sequential affinity chromatography steps to give Sp1 of an estimated 90% homogeneity with 30% yield. In addition, the use of tandem affinity columns containing different protein binding sites allows the simultaneous purification of multiple DNA binding proteins from the same extract. This method provides a means for the purification of rare sequence-specific DNA binding proteins, such as Sp1 and CAAT-binding transcription factor.

  17. High affinity ligands from in vitro selection: Complex targets

    PubMed Central

    Morris, Kevin N.; Jensen, Kirk B.; Julin, Carol M.; Weil, Michael; Gold, Larry

    1998-01-01

    Human red blood cell membranes were used as a model system to determine if the systematic evolution of ligands by exponential enrichment (SELEX) methodology, an in vitro protocol for isolating high-affinity oligonucleotides that bind specifically to virtually any single protein, could be used with a complex mixture of potential targets. Ligands to multiple targets were generated simultaneously during the selection process, and the binding affinities of these ligands for their targets are comparable to those found in similar experiments against pure targets. A secondary selection scheme, deconvolution-SELEX, facilitates rapid isolation of the ligands to targets of special interest within the mixture. SELEX provides high-affinity compounds for multiple targets in a mixture and might allow a means for dissecting complex biological systems. PMID:9501188

  18. 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.

  19. 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 of oxygen is computed to be 1.287 eV, at the full CI level using a 6s5p3d 2f Slater-type orbital basis and correlating only the 2p electrons. The best CASSCF-MRCI result including only 2p correlation is 1.263 eV. However, inclusion of 2s intrashell and 2s2p intershell coorelation increases the computed EA to 1.290 eV 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. The higher excitation contribution to the electron affinity is found to increase substantially with basis set completeness, especially when the 2s electrons are correlated. Relativistic effects are shown to make a small (less than 0.01 eV) change in the EA.

  20. Latest technologies for the enhancement of antibody affinity.

    PubMed

    Wark, Kim L; Hudson, Peter J

    2006-08-01

    High affinity antibodies are crucial both for the discovery and validation of biomarkers for human health and disease and as clinical diagnostic and therapeutic reagents. This review describes some of the latest technologies for the design, mutation and selection of high affinity antibodies that provide a paradigm for molecular evolution of a far wider range of proteins including enzymes. Strategies include both in vivo and in vitro methods and embrace the latest concepts for antibody display and selection. Specifically, affinity enhancement can be tailored to the target-binding surface, typically the complementary determining region (CDR) loops in antibodies, whereas enhanced stability, expression or catalytic properties can be affected by selected changes to the core protein scaffold. Together, these technologies provide a rapid and powerful strategy to drive the next generation of protein-based reagents for numerous clinical, environmental and agribusiness applications.

  1. 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.

  2. Considering affinity: an ethereal conversation (part two of three).

    PubMed

    Winsor, Mary P

    2015-06-01

    In 1840 Hugh Strickland published a diagram showing the relationships of genera of birds in the kingfisher family. Three years later he applied this mapping idea to genera of birds of prey and songbirds, creating a large wall chart that he displayed to colleagues but never published. Both of his diagrams featured a scale of degrees of affinity. The meaning of taxonomic affinity was something Darwin thought about deeply. Details in the chart undermine Strickland's claim that his method was purely inductive. PMID:25547607

  3. A quantum affine algebra for the deformed Hubbard chain

    NASA Astrophysics Data System (ADS)

    Beisert, Niklas; Galleas, Wellington; Matsumoto, Takuya

    2012-09-01

    The integrable structure of the one-dimensional Hubbard model is based on Shastry's R-matrix and the Yangian of a centrally extended \\mathfrak {sl}(2|2) superalgebra. Alcaraz and Bariev have shown that the model admits an integrable deformation whose R-matrix has recently been found. This R-matrix is of trigonometric type and here we derive its underlying exceptional quantum affine algebra. We also show how the algebra reduces to the above-mentioned Yangian and to the conventional quantum affine \\mathfrak {sl}(2|2) algebra in two special limits.

  4. 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.

  5. 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.

  6. 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.

  7. 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.

  8. 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

  9. 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

  10. "The Hunger Games": Literature, Literacy, and Online Affinity Spaces

    ERIC Educational Resources Information Center

    Curwood, Jen Scott

    2013-01-01

    This article examines adolescent literacy practices related to "The Hunger Games," a young adult novel and the first of a trilogy. By focusing on the interaction of social identities, discourses, and media paratexts within an online affinity space, this ethnographic study offers insight into how young adults engage with contemporary…

  11. 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.

  12. 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

  13. 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"…

  14. 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…

  15. 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…

  16. 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

  17. 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

  18. Feature Matching with Affine-Function Transformation Models.

    PubMed

    Li, Hongsheng; Huang, Xiaolei; Huang, Junzhou; Zhang, Shaoting

    2014-12-01

    Feature matching is an important problem and has extensive uses in computer vision. However, existing feature matching methods support either a specific or a small set of transformation models. In this paper, we propose a unified feature matching framework which supports a large family of transformation models. We call the family of transformation models the affine-function family, in which all transformations can be expressed by affine functions with convex constraints. In this framework, the goal is to recover transformation parameters for every feature point in a template point set to calculate their optimal matching positions in an input image. Given pairwise feature dissimilarity values between all points in the template set and the input image, we create a convex dissimilarity function for each template point. Composition of such convex functions with any transformation model in the affine-function family is shown to have an equivalent convex optimization form that can be optimized efficiently. Four example transformation models in the affine-function family are introduced to show the flexibility of our proposed framework. Our framework achieves 0.0 percent matching errors for both CMU House and Hotel sequences following the experimental setup in [6]. PMID:26353148

  19. Smooth surfaces from bilinear patches: Discrete affine minimal surfaces.

    PubMed

    Käferböck, Florian; Pottmann, Helmut

    2013-06-01

    Motivated by applications in freeform architecture, we study surfaces which are composed of smoothly joined bilinear patches. These surfaces turn out to be discrete versions of negatively curved affine minimal surfaces and share many properties with their classical smooth counterparts. We present computational design approaches and study special cases which should be interesting for the architectural application.

  20. Properties of an affine transport equation and its holonomy

    NASA Astrophysics Data System (ADS)

    Vines, Justin; Nichols, David A.

    2016-10-01

    An affine transport equation was used recently to study properties of angular momentum and gravitational-wave memory effects in general relativity. In this paper, we investigate local properties of this transport equation in greater detail. Associated with this transport equation is a map between the tangent spaces at two points on a curve. This map consists of a homogeneous (linear) part given by the parallel transport map along the curve plus an inhomogeneous part, which is related to the development of a curve in a manifold into an affine tangent space. For closed curves, the affine transport equation defines a "generalized holonomy" that takes the form of an affine map on the tangent space. We explore the local properties of this generalized holonomy by using covariant bitensor methods to compute the generalized holonomy around geodesic polygon loops. We focus on triangles and "parallelogramoids" with sides formed from geodesic segments. For small loops, we recover the well-known result for the leading-order linear holonomy (˜ Riemann × area), and we derive the leading-order inhomogeneous part of the generalized holonomy (˜ Riemann × area^{3/2}). Our bitensor methods let us naturally compute higher-order corrections to these leading results. These corrections reveal the form of the finite-size effects that enter into the holonomy for larger loops; they could also provide quantitative errors on the leading-order results for finite loops.

  1. 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

  2. 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

  3. 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.

  4. 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…

  5. Calculation of antibody affinity in homogeneous and heterogeneous systems.

    PubMed

    Chalquest, R R

    1988-12-01

    Antibody affinity is an important determinant of all antibody-antigen reactions. A new computer program, AFCRV, was developed to calculate binding constants with data from a radioimmunoassay on most microcomputers in the laboratory by using constant-ratio dilution curves. Evaluation of a homogeneous or heterogeneous antibody in the presence of a single antigen can be accomplished.

  6. 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

  7. 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.

  8. Semiempirical Theories of the Affinities of Negative Atomic Ions

    NASA Technical Reports Server (NTRS)

    Edie, John W.

    1961-01-01

    The determination of the electron affinities of negative atomic ions by means of direct experimental investigation is limited. To supplement the meager experimental results, several semiempirical theories have been advanced. One commonly used technique involves extrapolating the electron affinities along the isoelectronic sequences, The most recent of these extrapolations Is studied by extending the method to Include one more member of the isoelectronic sequence, When the results show that this extension does not increase the accuracy of the calculations, several possible explanations for this situation are explored. A different approach to the problem is suggested by the regularities appearing in the electron affinities. Noting that the regular linear pattern that exists for the ionization potentials of the p electrons as a function of Z, repeats itself for different degrees of ionization q, the slopes and intercepts of these curves are extrapolated to the case of the negative Ion. The method is placed on a theoretical basis by calculating the Slater parameters as functions of q and n, the number of equivalent p-electrons. These functions are no more than quadratic in q and n. The electron affinities are calculated by extending the linear relations that exist for the neutral atoms and positive ions to the negative ions. The extrapolated. slopes are apparently correct, but the intercepts must be slightly altered to agree with experiment. For this purpose one or two experimental affinities (depending on the extrapolation method) are used in each of the two short periods. The two extrapolation methods used are: (A) an isoelectronic sequence extrapolation of the linear pattern as such; (B) the same extrapolation of a linearization of this pattern (configuration centers) combined with an extrapolation of the other terms of the ground configurations. The latter method Is preferable, since it requires only experimental point for each period. The results agree within

  9. 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

  10. 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

  11. Histology and affinity of the earliest armoured vertebrate.

    PubMed

    Sansom, Ivan J; Donoghue, Philip C J; Albanesi, Guillermo

    2005-12-22

    Arandaspids are the earliest skeletonizing vertebrates known from articulated remains. Despite a wealth of data, their affinity remains questionable because they exhibit a random mixture of primitive and derived characteristics. We constrain the affinity of arandaspids by providing the first detailed characterization of their dermoskeleton which is revealed to be three-layered, composed of a basal laminated, cancellous middle and tubercular superficial layers. All three layers are composed of acellular bone but the superficial layer also includes dentine and enameloid, comprising the tubercles. As such, the composition of the arandaspid dermoskeleton is common to heterostracans and astraspids, supporting existing hypotheses of early vertebrate phylogeny. This emphasizes the peculiarity of existing interpretations of aranadaspid anatomy and there is need for a complete reappraisal of the existing anatomical data.

  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. 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.

  14. Affinity screening using competitive binding with fluorine-19 hyperpolarized ligands.

    PubMed

    Kim, Yaewon; Hilty, Christian

    2015-04-13

    Fluorine-19 NMR and hyperpolarization form a powerful combination for drug screening. Under a competitive equilibrium with a selected fluorinated reporter ligand, the dissociation constant (K(D)) of other ligands of interest is measurable using a single-scan Carr-Purcell-Meiboom-Gill (CPMG) experiment, without the need for a titration. This method is demonstrated by characterizing the binding of three ligands with different affinities for the serine protease trypsin. Monte Carlo simulations show that the highest accuracy is obtained when about one-half of the bound reporter ligand is displaced in the binding competition. Such conditions can be achieved over a wide range of affinities, allowing for rapid screening of non-fluorinated compounds when a single fluorinated ligand for the binding pocket of interest is known.

  15. 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}.

  16. 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

  17. 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.

  18. 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

  19. Isotope shift in the electron affinity of lithium.

    PubMed

    Bubin, Sergiy; Komasa, Jacek; Stanke, Monika; Adamowicz, Ludwik

    2009-12-21

    Very accurate electron affinity (EA) calculations of (6)Li and (7)Li (and (infinity)Li) 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 (7)Li and (6)Li EAs are 4984.9842(30) and 4984.9015(30) cm(-1), respectively, and of the (7)Li/(6)Li EA isotope shift is 0.0827 cm(-1).

  20. Affinity enhancement of antibodies: how low-affinity antibodies produced early in immune responses are followed by high-affinity antibodies later and in memory B-cell responses.

    PubMed

    Eisen, Herman N

    2014-05-01

    The antibodies produced initially in response to most antigens are high molecular weight (MW) immunoglobulins (IgM) with low affinity for the antigen, while the antibodies produced later are lower MW classes (e.g., IgG and IgA) with, on average, orders of magnitude higher affinity for that antigen. These changes, often termed affinity maturation, take place largely in small B-cell clusters (germinal center; GC) in lymphoid tissues in which proliferating antigen-stimulated B cells express the highly mutagenic cytidine deaminase that mediates immunoglobulin class-switching and sequence diversification of the immunoglobulin variable domains of antigen-binding receptors on B cells (BCR). Of the large library of BCR-mutated B cells thus rapidly generated, a small minority with affinity-enhancing mutations are selected to survive and differentiate into long-lived antibody-secreting plasma cells and memory B cells. BCRs are also endocytic receptors; they internalize and cleave BCR-bound antigen, yielding peptide-MHC complexes that are recognized by follicular helper T cells. Imperfect correlation between BCR affinity for antigen and cognate T-cell engagement may account for the increasing affinity heterogeneity that accompanies the increasing average affinity of antibodies. Conservation of mechanisms underlying mutation and selection of high-affinity antibodies over the ≈200 million years of evolution separating bird and mammal lineages points to the crucial role of antibody affinity enhancement in adaptive immunity.

  1. High affinity DNA-microtubule associated protein interaction.

    PubMed

    Marx, K A

    1992-07-01

    We have isolated the MAP/tau proteins from twice-cycled chick brain microtubule preparations and demonstrated that they are responsible for the nitrocellulose DNA binding activity we and others have measured. Using the isolated MAP/tau proteins we then measured the apparent affinity constant K(app) for the homologous chick DNA interaction and found evidence for two equilibrium affinity classes-a K(app) = 6 x 10(7) M-1, responsible for the bulk of the DNA binding activity and a small (less than 10%) higher affinity K(app) = 10(8) - 10(9) M-1, likely due to sequence specific binding protein species. Using the same chick brain MAP-tau protein, a heterologous interaction with D. melanogaster DNA, was found to possess just the lower affinity class-K(app) = 2 x 10(7) M-1. Under stringent binding conditions we carried out equilibrium nitrocellulose filter binding experiments in a ternary reaction mixture at constant MAP/tau protein and 35S radiolabelled chick DNA concentration using increasing and excess concentrations of competitor DNAs of different sources. The order of competitor strengths found was-chick DNA greater than mouse DNA greater than D. melanogaster = E. coli. DNA. These data and specifically the homologous DNA: protein case being the strongest competitor corroborate our previous studies using total microtubule protein and provide new evidence for a conserved interaction of a small DNA sequence class with MAP/tau protein species. Moreover, these data allow us to conclude that the conserved DNA sequence: MAP/tau protein interactions do not critically depend upon any energetic feature co-involving tubulin for their properties since tubulin is absent from these preparations.

  2. Chlorophenylpiperazine analogues as high affinity dopamine transporter ligands.

    PubMed

    Motel, William C; Healy, Jason R; Viard, Eddy; Pouw, Buddy; Martin, Kelly E; Matsumoto, Rae R; Coop, Andrew

    2013-12-15

    Selective σ2 ligands continue to be an active target for medications to attenuate the effects of psychostimulants. In the course of our studies to determine the optimal substituents in the σ2-selective phenyl piperazines analogues with reduced activity at other neurotransmitter systems, we discovered that 1-(3-chlorophenyl)-4-phenethylpiperazine actually had preferentially increased affinity for dopamine transporters (DAT), yielding a highly selective DAT ligand. PMID:24211020

  3. Affinity Purification of Protein Complexes Using TAP Tags

    PubMed Central

    Gerace, Erica; Moazed, Danesh

    2016-01-01

    This protocol is used for the isolation and analysis of protein complexes using the tandem affinity purification (TAP) tag system. The protocol describes the purification of a protein fused to a TAP tag comprised of two protein A domains and the calmodulin binding peptide separated by a TEV cleavage site. This is a powerful technique for rapid purification of protein complexes and the analysis of their stoichiometric composition, posttranslational modifications, structure, and functional activities. PMID:26096502

  4. Selective high affinity polydentate ligands and methods of making such

    DOEpatents

    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.

  5. 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

  6. Affine transformations capture beak shape variation in Darwin's Finches

    NASA Astrophysics Data System (ADS)

    Brenner, Michael; Campas, Otger; Mallarino, Riccardo; Abzhanov, Arhat

    2009-11-01

    Evolution by natural selection has resulted in extraordinary morphological complexity of living organisms, whose description has thus far defied any precise mathematical characterization linked to the underlying developmental genetics. Here we demonstrate that the morphological diversity of the beaks of Darwin's finches, the classical example of adaptive morphological radiation, is quantitatively accounted for through the mathematical group of affine transformations. Specifically, we show that all beak shapes of Ground Finches (genus Geospiza) are related by scaling transformations (a subgroup of the affine group), and the same scheme occurs for all the beak shapes of Tree and Warbler finches. This analysis shows that the beak shapes within each of these groups differ only by their scales, such as length and depth, each of which is knownto be under genetic control.The complete morphological variability within the beaks of Darwin's finches can be explained by extending the scaling transformations to the entire affine group, by including shear transformations. Altogether our results suggest that the mathematical theory of groups can help decode morphological variability, and points to a potentially hierarchical structure of morphological diversity and the underlying developmental processes.

  7. Synthesis and NMDA receptor affinity of fluorinated dioxadrol analogues.

    PubMed

    Banerjee, Ashutosh; Schepmann, Dirk; Wünsch, Bernhard

    2010-06-01

    A series of dioxadrol analogues with fluorine substituents in position 4 of the piperidine ring has been synthesized and pharmacologically evaluated. The key step in the synthesis was the fluorination of diastereomeric piperidones 6a and 6c as well as diastereomeric alcohols 9a and 9c with DAST. The reaction of the alcohols 9a and 9c took place with inversion of configuration. After removal of the Cbz-protective group, the NMDA receptor affinities of the resulting secondary amines 8a, 8c, 12b, and 12d were investigated in receptor binding studies. It was shown that the like-configuration of the ring junction was crucial for high NMDA receptor affinity. An axially oriented fluorine atom in position 4 led to 2-(2,2-diphenyl-1,3-dioxolan-4-yl)-4-fluoropiperidine (12d, WMS-2517) with a K(i)-value of 27nM. The NMDA receptor affinity of 8c (WMS-2513) with an additional fluorine atom in equatorial 4-position was slightly reduced (K(i)=81 nM). Both fluorinated dioxadrol derivatives 8c and 12d showed high selectivity against sigma(1) and sigma(2) receptors as well as the polyamine binding site of NR2B receptors.

  8. 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

  9. 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

  10. 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

  11. 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

  12. 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

  13. 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

  14. 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

  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.

  16. 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.

  17. Multiple lectin detection by cell membrane affinity binding.

    PubMed

    Ribeiro, Ana; Catarino, Sofia; Ferreira, Ricardo Boavida

    2012-05-01

    Assuming that lectins evolved to recognise relatively complex and branched oligosaccharides or parts of them, rather than simple sugars, a procedure based on lectin affinity binding to isolated erythrocyte (or any other cell type) membranes is proposed. This methodology was validated using six pure commercial lectins, as well as lectins from total protein extracts of Arbutus unedo leaves. All commercial lectins, as well as five polypeptides from A. unedo leaves bound to the glycosylated membrane receptors and were eluted by the corresponding sugars. When compared to the standard affinity chromatography procedure involving an individual sugar bound to a solid matrix, the new method provides a single-step, effective detection method for lectins and allows the rapid screening of their profile present in any unknown protein solution, indicates their biological carbohydrate affinities as well as their sugar specificities (if any), enables the simultaneous analysis of a large number of samples, does not require any pre-purification steps, permits detection of additional lectins and provides data which are more relevant from the physiological point of view. PMID:22381939

  18. 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

  19. Relative binding affinities of monolignols to horseradish peroxidase

    DOE PAGES

    Sangha, Amandeep K.; Petridis, Loukas; Cheng, Xiaolin; Smith, Jeremy C.

    2016-07-22

    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 andmore » 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. Furthermore, 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.« less

  20. Molecular fine-tuning of affinity maturation in germinal centers.

    PubMed

    Wu, Yee Ling; Rada, Cristina

    2016-01-01

    The development of high-affinity antibodies in response to infection is an iterative process in which B cells cycle between proliferation/somatic hypermutation and antigen-driven selection. These processes occur within specific regions of the secondary lymphoid structures known as germinal centers (GCs) and the environmental and signaling cues provided by these regions guide the GC reactions that drive B cell maturation and antibody production, ultimately determining B cell fate. In this issue of the JCI, Nakagawa and colleagues examine the role of miR-155, a microRNA that is required for GC development and the production of high-affinity antibodies. They show that miR-155 is highly expressed in positively selected B cells and promotes survival of these cells by orienting the Myc transcription program toward survival rather than apoptosis through the inhibition of the transcriptional regulator JARID2. These findings illustrate the fine balance between apoptosis and proliferation that is required for the development of high-affinity antibodies.

  1. 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

  2. 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)

  3. 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

  4. 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.

  5. 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.

  6. 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

  7. Recent improvements to Binding MOAD: a resource for protein–ligand binding affinities and structures

    PubMed Central

    Ahmed, Aqeel; Smith, Richard D.; Clark, Jordan J.; Dunbar, James B.; Carlson, Heather A.

    2015-01-01

    For over 10 years, Binding MOAD (Mother of All Databases; http://www.BindingMOAD.org) has been one of the largest resources for high-quality protein–ligand complexes and associated binding affinity data. Binding MOAD has grown at the rate of 1994 complexes per year, on average. Currently, it contains 23 269 complexes and 8156 binding affinities. Our annual updates curate the data using a semi-automated literature search of the references cited within the PDB file, and we have recently upgraded our website and added new features and functionalities to better serve Binding MOAD users. In order to eliminate the legacy application server of the old platform and to accommodate new changes, the website has been completely rewritten in the LAMP (Linux, Apache, MySQL and PHP) environment. The improved user interface incorporates current third-party plugins for better visualization of protein and ligand molecules, and it provides features like sorting, filtering and filtered downloads. In addition to the field-based searching, Binding MOAD now can be searched by structural queries based on the ligand. In order to remove redundancy, Binding MOAD records are clustered in different families based on 90% sequence identity. The new Binding MOAD, with the upgraded platform, features and functionalities, is now equipped to better serve its users. PMID:25378330

  8. Advances and applications of binding affinity prediction methods in drug discovery.

    PubMed

    Parenti, Marco Daniele; Rastelli, Giulio

    2012-01-01

    Nowadays, the improvement of R&D productivity is the primary commitment in pharmaceutical research, both in big pharma and smaller biotech companies. To reduce costs, to speed up the discovery process and to increase the chance of success, advanced methods of rational drug design are very helpful, as demonstrated by several successful applications. Among these, computational methods able to predict the binding affinity of small molecules to specific biological targets are of special interest because they can accelerate the discovery of new hit compounds. Here we provide an overview of the most widely used methods in the field of binding affinity prediction, as well as of our own work in developing BEAR, an innovative methodology specifically devised to overtake some limitations in existing approaches. The BEAR method was successfully validated against different biological targets, and proved its efficacy in retrieving active compounds from virtual screening campaigns. The results obtained so far indicate that BEAR may become a leading tool in the drug discovery pipeline. We primarily discuss advantages and drawbacks of each technique and show relevant examples and applications in drug discovery.

  9. 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.

  10. 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

  11. Minimization of the root of a quadratic functional under a system of affine equality constraints with application to portfolio management

    NASA Astrophysics Data System (ADS)

    Landsman, Zinoviy

    2008-10-01

    We present an explicit closed form solution of the problem of minimizing the root of a quadratic functional subject to a system of affine constraints. The result generalizes Z. Landsman, Minimization of the root of a quadratic functional under an affine equality constraint, J. Comput. Appl. Math. 2007, to appear, see , articles in press, where the optimization problem was solved under only one linear constraint. This is of interest for solving significant problems pertaining to financial economics as well as some classes of feasibility and optimization problems which frequently occur in tomography and other fields. The results are illustrated in the problem of optimal portfolio selection and the particular case when the expected return of finance portfolio is certain is discussed.

  12. Improving affinity chromatography resin efficiency using semi-continuous chromatography.

    PubMed

    Mahajan, Ekta; George, Anupa; Wolk, Bradley

    2012-03-01

    Protein A affinity chromatography is widely used for purification of monoclonal antibodies (MAbs) from harvested cell culture fluid (HCCF). At the manufacturing scale, the HCCF is typically loaded on a single Protein A affinity chromatography column in cycles until all of the HCCF is processed. Protein A resin costs are significant, comprising a substantial portion of the raw material costs in MAb manufacturing. Cost can be reduced by operating the process continuously using multiple smaller columns to a higher binding capacity in lieu of one industrial scale column. In this study, a series of experiments were performed using three 1-ml Hi-Trap™ MabSelect SuRe™ columns on a modified ÄKTA™ system operated according to the three Column Periodic Counter Current Chromatography (3C PCC) principle. The columns were loaded individually at different times until the 70% breakthrough point was achieved. The HCCF with unbound protein from the column was then loaded onto the next column to capture the MAb, preventing any protein loss. At any given point, all three columns were in operation, either loading or washing, enabling a reduction in processing time. The product yield and quality were evaluated and compared with a batch process to determine the effect of using the three column continuous process. The continuous operation shows the potential to reduce both resin volume and buffer consumption by ∼40%, however the system hardware and the process is more complex than the batch process. Alternative methods using a single standard affinity column, such as recycling load effluent back to the tank or increasing residence time, were also evaluated to improve Protein A resin efficiency. These alternative methods showed similar cost benefits but required longer processing time. PMID:22265178

  13. 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

  14. Computational study of coagulation factor VIIa's affinity for phospholipid membranes.

    PubMed

    Taboureau, Olivier; Olsen, Ole Hvilsted

    2007-02-01

    The interaction between the gamma-carboxyglutamic acid-rich domain of coagulation factor VIIa (FVIIa), a vitamin-K-dependent enzyme, and phospholipid membranes plays a major role in initiation of blood coagulation. However, despite a high sequence and structural similarity to the Gla domain of other vitamin-K-dependent enzymes with a high membrane affinity, its affinity for negatively charged phospholipids is poor. A few amino acid differences are responsible for this observation. Based on the X-ray structure of lysophosphatidylserine (lysoPS) bound to the Gla domain of bovine prothrombin (Prth), models of the Gla domain of wildtype FVIIa and mutated FVIIa Gla domains in complex with lysoPS were built. Molecular dynamics (MD) and steered molecular dynamics (SMD) simulations on the complexes were applied to investigate the significant difference in the binding affinity. The MD simulation approach provides a structural and dynamic support to the role of P10Q and K32E mutations in the improvement of the membrane contact. Hence, rotation of the Gly11 main chain generated during the MD simulation results in a hydrogen bond with Q10 side chain as well as the appearance of a hydrogen bond between E32 and Q10 forcing the loop harbouring Arg9 and Arg15 to shrink and thereby enhances the accessibility of the phospholipids to the calcium ions. Furthermore, the application of the SMD simulation method to dissociate C6-lysoPS from a series of Gla domain models exhibits a ranking of the rupture force that can be useful in the interpretation of the PS interaction with Gla domains. Finally, adiabatic mapping of Gla6 residue in FVIIa with or without insertion of Tyr4 confirms the critical role of the insertion on the conformation of the side chain Gla6 in FVIIa and the corresponding Gla7 in Prth. PMID:17131117

  15. 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

  16. Biospecific affinity chromatographic purification of octopine dehydrogenase from molluscs.

    PubMed

    Mulcahy, P; Griffin, T; O'Carra, P

    1997-02-01

    The development of a biospecific affinity chromatographic method for the purification of octopine dehydrogenase from molluscs is described. The method utilizes immobilized NAD+ derivatives in conjunction with soluble specific substrates to promote binding. Using this method, octopine dehydrogenase has been purified to electrophoretic homogeneity in a single chromatographic step from three different marine invertebrate sources [the queen scallop, Chlamys opercularis (adductor muscle), the great scallop, Pecten maximus (adductor muscle), and the squid Loligo vulgaris (mantle muscle)]. However, the system is not applicable to the purification of octopine dehydrogenase from some other marine invertebrate sources investigated (the mussel Mytilus edulis and the topshell Monodonta lineata). PMID:9116492

  17. Isolation of human lactate dehydrogenase isoenzyme X by affinity chromatography.

    PubMed Central

    Kolk, A H; van Kuyk, L; Boettcher, B

    1978-01-01

    Human isoenzyme LDH-X (lactate dehydrogenase isoenzyme X) was isolated from seminal fluid of frozen semen samples by affinity chromatography by using oxamate-Sepharose and AMP-Sepharose. In the presence of 1.6 mM-NAD+, isoenzyme LDH-X does not bind to AMP-Sepharose, whereas the other lactate dehydrogenase isoenzymes do. This is the crucial point in the isolation of isoenzyme LDH-X from the other isoenzymes. The purified human isoenzyme LDH-X had a specific activity of 146 units/mg of protein. Images Fig. 2. Fig. 3. PMID:213050

  18. High affinity of water-soluble cryptophanes for cesium cations.

    PubMed

    Brotin, Thierry; Montserret, Roland; Bouchet, Aude; Cavagnat, Dominique; Linares, Mathieu; Buffeteau, Thierry

    2012-01-20

    Exceptionally high affinity for cesium cations was achieved in aqueous solution using two enantiopure cryptophanes. Complexation of cesium was evidenced by (133)Cs NMR spectroscopy and by electronic circular dichroism (ECD). Binding constants as high as 6 × 10(9) M(-1) have been measured by isothermal titration calorimetry (ITC). Very strong complexation of rubidium cations (K ~10(6) M(-1)) has also been measured. Chiral hosts allowed the detection of the two cations at low concentrations (μM) using ECD.

  19. Affinity enhancement by dendritic side chains in synthetic carbohydrate receptors.

    PubMed

    Destecroix, Harry; Renney, Charles M; Mooibroek, Tiddo J; Carter, Tom S; Stewart, Patrick F N; Crump, Matthew P; Davis, Anthony P

    2015-02-01

    Dendritic side chains have been used to modify the binding environment in anthracene-based synthetic carbohydrate receptors. Control of length, charge, and branching enabled the positioning of side-chain carboxylate groups in such a way that they assisted in binding substrates rather than blocking the cavity. Conformational degeneracy in the dendrimers resulted in effective preorganization despite the flexibility of the system. Strong binding was observed to glucosammonium ions in water, with Ka values up to 7000 M(-1) . Affinities for uncharged substrates (glucose and N-acetylglucosamine) were also enhanced, despite competition from solvent and the absence of electrostatic interactions. PMID:25645064

  20. Friction and plasticity between self-affine surfaces

    NASA Astrophysics Data System (ADS)

    Luan, Binquan; Robbins, Mark; Harrison, Judith

    2006-03-01

    Simulations are used to study the contact area and adhesion between two amorphous solids with self-affine fractal surfaces, and the results are compared to continuum calculations. The friction between non-adhesive surfaces is proportional to load, but the coefficient of friction increases with roughness. The friction is much higher than expected for elasticallly deforming surfaces,^* and substantial plastic deformation is observed. Indeed, friction forces for different surface roughness collapse when plotted against the number of plastic rearrangements per unit sliding distance. Including adhesion leads to an increase in both friction and plasticity. ^* M. H. Müser, L. Wenning, and M. O. Robbins, Phys. Rev. Lett. 86, 1295 (2001).

  1. Nine switch-affine neurons suffice for Turing universality.

    PubMed

    Siegelmann, H T.; Margenstern, M

    1999-06-01

    In a previous work Pollack showed that a particular type of heterogeneous processor network is Turing universal. Siegelmann and Sontag (1991) showed the universality of homogeneous networks of first-order neurons having piecewise-linear activation functions. Their result was generalized by Kilian and Siegelmann (1996) to include various sigmoidal activation functions. Here we focus on a type of high-order neurons called switch-affine neurons, with piecewise-linear activation functions, and prove that nine such neurons suffice for simulating universal Turing machines. PMID:12662670

  2. An affine projection algorithm using grouping selection of input vectors

    NASA Astrophysics Data System (ADS)

    Shin, JaeWook; Kong, NamWoong; Park, PooGyeon

    2011-10-01

    This paper present an affine projection algorithm (APA) using grouping selection of input vectors. To improve the performance of conventional APA, the proposed algorithm adjusts the number of the input vectors using two procedures: grouping procedure and selection procedure. In grouping procedure, the some input vectors that have overlapping information for update is grouped using normalized inner product. Then, few input vectors that have enough information for for coefficient update is selected using steady-state mean square error (MSE) in selection procedure. Finally, the filter coefficients update using selected input vectors. The experimental results show that the proposed algorithm has small steady-state estimation errors comparing with the existing algorithms.

  3. Calcium affinity of human α-actinin 1.

    PubMed

    Backman, Lars

    2015-01-01

    Due to alternative splicing, the human ACTN1 gene codes for three different transcripts of α-actinin; one isoform that is expressed only in the brain and two with a more general expression pattern. The sequence difference is located to the C-terminal domains and the EF-hand motifs. Therefore, any functional or structural distinction should involve this part of the protein. To investigate this further, the calcium affinities of these three isoforms of α-actinin 1 have been determined by isothermal calorimetry. PMID:26020004

  4. Kinetic analysis of drug-protein interactions by affinity chromatography.

    PubMed

    Bi, Cong; Beeram, Sandya; Li, Zhao; Zheng, Xiwei; Hage, David S

    2015-10-01

    Information on the kinetics of drug-protein interactions is of crucial importance in drug discovery and development. Several methods based on affinity chromatography have been developed in recent years to examine the association and dissociation rates of these processes. These techniques include band-broadening measurements, the peak decay method, peak fitting methods, the split-peak method, and free fraction analysis. This review will examine the general principles and applications of these approaches and discuss their use in the characterization, screening and analysis of drug-protein interactions in the body. PMID:26724332

  5. Enrichment of Phosphopeptides via Immobilized Metal Affinity Chromatography.

    PubMed

    Swaney, Danielle L; Villén, Judit

    2016-03-01

    Immobilized metal affinity chromatography (IMAC) is a frequently used method for the enrichment of phosphorylated peptides from complex, cellular lysate-derived peptide mixtures. Here we outline an IMAC protocol that uses iron-chelated magnetic beads to selectively isolate phosphorylated peptides for mass spectrometry-based proteomic analysis. Under acidic conditions, negatively charged phosphoryl modifications preferentially bind to positively charged metal ions (e.g., Fe(3+), Ga(3+)) on the beads. After washing away nonphosphorylated peptides, a pH shift to basic conditions causes the elution of bound phosphopeptides from the metal ion. Under optimal conditions, very high specificity for phosphopeptides can be achieved. PMID:26933247

  6. Analyzing a kinetic titration series using affinity biosensors.

    PubMed

    Karlsson, Robert; Katsamba, Phinikoula S; Nordin, Helena; Pol, Ewa; Myszka, David G

    2006-02-01

    The classical method of measuring binding constants with affinity-based biosensors involves testing several analyte concentrations over the same ligand surface and regenerating the surface between binding cycles. Here we describe an alternative approach to collecting kinetic binding data, which we call "kinetic titration." This method involves sequentially injecting an analyte concentration series without any regeneration steps. Through a combination of simulation and experimentation, we show that this method can be as robust as the classical method of analysis. In addition, kinetic titrations can be more efficient than the conventional data collection method and allow us to fully characterize analyte binding to ligand surfaces that are difficult to regenerate.

  7. Representations of affine superalgebras and mock theta functions. III

    NASA Astrophysics Data System (ADS)

    Kac, V. G.; Wakimoto, M.

    2016-08-01

    We study modular invariance of normalized supercharacters of tame integrable modules over an affine Lie superalgebra, associated to an arbitrary basic Lie superalgebra g. For this we develop a several step modification process of multivariable mock theta functions, where at each step a Zwegers' type 'modifier' is used. We show that the span of the resulting modified normalized supercharacters is \\operatorname{SL}_2( Z)-invariant, with the transformation matrix equal, in the case the Killing form on g is non-degenerate, to that for the basic defect 0 subalgebra g^! of g, orthogonal to a maximal isotropic set of roots of g.

  8. [Progresses in screening active compounds from herbal medicine by affinity chromatography].

    PubMed

    Feng, Ying-shu; Tong, Shan-shan; Xu, Xi-ming; Yu, Jiang-nan

    2015-03-01

    Affinity chromatography is a chromatographic method for separating molecules using the binding characteristics of the stationary phase with potential drug molecules. This method can be performed as a high throughput screening method and a chromatographic separation method to screen a variety of active drugs. This paper summarizes the history of affinity chromatography, screening technology of affinity chromatography, and application of affinity chromatography in screening bio-active compounds in herbal medicines, and then discusses its application prospects, in order to broaden applications of the affinity chromatography in drug screening. PMID:26226740

  9. Biocompatible silicon surfaces through orthogonal click chemistries and a high affinity silicon oxide binding peptide.

    PubMed

    Hassert, Rayk; Pagel, Mareen; Ming, Zhou; Häupl, Tilmann; Abel, Bernd; Braun, Klaus; Wiessler, Manfred; Beck-Sickinger, Annette G

    2012-10-17

    Multifunctionality is gaining more and more importance in the field of improved biomaterials. Especially peptides feature a broad chemical variability and are versatile mediators between inorganic surfaces and living cells. Here, we synthesized a unique peptide that binds to SiO(2) with nM affinity. We equipped the peptide with the bioactive integrin binding c[RGDfK]-ligand and a fluorescent probe by stepwise Diels-Alder reaction with inverse electron demand and copper(I) catalyzed azide-alkyne cycloaddition. For the first time, we report the generation of a multifunctional peptide by combining these innovative coupling reactions. The resulting peptide displayed an outstanding binding to silicon oxide and induced a significant increase in cell spreading and cell viability of osteoblasts on the oxidized silicon surface.

  10. Sequential class switching is required for the generation of high affinity IgE antibodies

    PubMed Central

    Xiong, Huizhong; Dolpady, Jayashree; Wabl, Matthias; Curotto de Lafaille, Maria A.

    2012-01-01

    IgE antibodies with high affinity for their antigens can be stably cross-linked at low concentrations by trace amounts of antigen, whereas IgE antibodies with low affinity bind their antigens weakly. In this study, we find that there are two distinct pathways to generate high and low affinity IgE. High affinity IgE is generated through sequential class switching (μ→γ→ε) in which an intermediary IgG phase is necessary for the affinity maturation of the IgE response, where the IgE inherits somatic hypermutations and high affinity from the IgG1 phase. In contrast, low affinity IgE is generated through direct class switching (μ→ε) and is much less mutated. Mice deficient in IgG1 production cannot produce high affinity IgE, even after repeated immunizations. We demonstrate that a small amount of high affinity IgE can cause anaphylaxis and is pathogenic. Low affinity IgE competes with high affinity IgE for binding to Fcε receptors and prevents anaphylaxis and is thus beneficial. PMID:22249450

  11. Challenges and recent advances in affinity purification of tag-free proteins.

    PubMed

    Guan, Dongli; Chen, Zhilei

    2014-07-01

    There is currently no generic, simple, lowcost method for affinity chromatographic purification of proteins in which the purified product is free of appended tags. Existing approaches for the purification of tagless proteins fall into two broad categories: (1) direct affinity-based capture of tag-free proteins that utilize affinity ligands specific to the target protein or class of target protein, and (2) removal of an appended affinity tag following tag-mediated protein capture. This paper reviews current state-of-the-art approaches for tagless protein purification in both categories, including specific examples of affinity ligands used for the capture of different classes of proteins and cleavage systems for affinity tag removal following chromatographic capture. A particular focus of this review is on recent developments in affinity tag removal systems utilizing split inteins. PMID:24658742

  12. 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

  13. Affinity sensor using 3-aminophenylboronic acid for bacteria detection.

    PubMed

    Wannapob, Rodtichoti; Kanatharana, Proespichaya; Limbut, Warakorn; Numnuam, Apon; Asawatreratanakul, Punnee; Thammakhet, Chongdee; Thavarungkul, Panote

    2010-10-15

    Boronic acid that can reversibly bind to diols was used to detect bacteria through its affinity binding reaction with diol-groups on bacterial cell walls. 3-aminophenylboronic acid (3-APBA) was immobilized on a gold electrode via a self-assembled monolayer. The change in capacitance of the sensing surface caused by the binding between 3-APBA and bacteria in a flow system was detected by a potentiostatic step method. Under optimal conditions the linear range of 1.5×10(2)-1.5×10(6) CFU ml(-1) and the detection limit of 1.0×10(2) CFU ml(-1) was obtained. The sensing surface can be regenerated and reused up to 58 times. The method was used for the analysis of bacteria in several types of water, i.e., bottled, well, tap, reservoir and wastewater. Compared with the standard plate count method, the results were within one standard deviation of each other. The proposed method can save both time and cost of analysis. The electrode modified with 3-APBA would also be applicable to the detection of other cis-diol-containing analytes. The concept could be extended to other chemoselective ligands, offering less expensive and more robust affinity sensors for a wide range of compounds.

  14. Influence of affinity on antibody determination in microtiter ELISA systems

    SciTech Connect

    Peterman, J.H.; Voss, E.W. Jr.; Butler, J.E.

    1986-03-01

    Theoretically, all immunoassays are affinity (Ka) dependent when the product of the antibody (Ab) Ka and the free epitope concentration is less than 10. Thus, the degree of dependence on Ka depends on the concentration of available antigen in the system. The authors examined the binding of /sup 125/I-anti-fluorescein (a-FLU) monoclonal antibodies of different affinities to FLU-gelatin adsorbed on Immunlon 2 microtiter plates. Data obtained were in general agreement with our theoretical predictions; the percent of /sup 125/I-a-FLU which bound correlated with Ka, as did the shape of the titration curves. Measurement of 5 a-FLU monoclonals by the ELISA showed that the determination of Ab concentrations depends on the FLU-gelatin concentration, epitope density, and on the relationship between the Kas of test samples and the reference standard Ab preparation. Thus the ELISA is Ka dependent and should not be used routinely to estimate the absolute amount to Ab in unknown samples. However, the Ka dependency of the ELISA might provide a convenient assay for the estimation of the relative functional Ka (rfKa) of antibody preparations.

  15. Telonemia, a new protist phylum with affinity to chromist lineages.

    PubMed

    Shalchian-Tabrizi, K; Eikrem, W; Klaveness, D; Vaulot, D; Minge, M A; Le Gall, F; Romari, K; Throndsen, J; Botnen, A; Massana, R; Thomsen, H A; Jakobsen, K S

    2006-07-22

    Recent molecular investigations of marine samples taken from different environments, including tropical, temperate and polar areas, as well as deep thermal vents, have revealed an unexpectedly high diversity of protists, some of them forming deep-branching clades within important lineages, such as the alveolates and heterokonts. Using the same approach on coastal samples, we have identified a novel group of protist small subunit (SSU) rDNA sequences that do not correspond to any phylogenetic group previously identified. Comparison with other sequences obtained from cultures of heterotrophic protists showed that the environmental sequences grouped together with Telonema, a genus known since 1913 but of uncertain taxonomic affinity. Phylogenetic analyses using four genes (SSU, Hsp90, alpha-tubulin and beta-tubulin), and accounting for gamma- and covarion-distributed substitution rates, revealed Telonema as a distinct group of species branching off close to chromist lineages. Consistent with these gene trees, Telonema possesses ultrastructures revealing both the distinctness of the group and the evolutionary affinity to chromist groups. Altogether, the data suggest that Telonema constitutes a new eukaryotic phylum, here defined as Telonemia, possibly representing a key clade for the understanding of the early evolution of bikont protist groups, such as the proposed chromalveolate supergroup. PMID:16790418

  16. Electron affinity of cubic boron nitride terminated with vanadium oxide

    NASA Astrophysics Data System (ADS)

    Yang, Yu; Sun, Tianyin; Shammas, Joseph; Kaur, Manpuneet; Hao, Mei; Nemanich, Robert J.

    2015-10-01

    A thermally stable negative electron affinity (NEA) for a cubic boron nitride (c-BN) surface with vanadium-oxide-termination is achieved, and its electronic structure was analyzed with in-situ photoelectron spectroscopy. The c-BN films were prepared by electron cyclotron resonance plasma-enhanced chemical vapor deposition employing BF3 and N2 as precursors. Vanadium layers of ˜0.1 and 0.5 nm thickness were deposited on the c-BN surface in an electron beam deposition system. Oxidation of the metal layer was achieved by an oxygen plasma treatment. After 650 °C thermal annealing, the vanadium oxide on the c-BN surface was determined to be VO2, and the surfaces were found to be thermally stable, exhibiting an NEA. In comparison, the oxygen-terminated c-BN surface, where B2O3 was detected, showed a positive electron affinity of ˜1.2 eV. The B2O3 evidently acts as a negatively charged layer introducing a surface dipole directed into the c-BN. Through the interaction of VO2 with the B2O3 layer, a B-O-V layer structure would contribute a dipole between the O and V layers with the positive side facing vacuum. The lower enthalpy of formation for B2O3 is favorable for the formation of the B-O-V layer structure, which provides a thermally stable surface dipole and an NEA surface.

  17. Reflection symmetry detection using locally affine invariant edge correspondence.

    PubMed

    Wang, Zhaozhong; Tang, Zesheng; Zhang, Xiao

    2015-04-01

    Reflection symmetry detection receives increasing attentions in recent years. The state-of-the-art algorithms mainly use the matching of intensity-based features (such as the SIFT) within a single image to find symmetry axes. This paper proposes a novel approach by establishing the correspondence of locally affine invariant edge-based features, which are superior to the intensity based in the aspects that it is insensitive to illumination variations, and applicable to textureless objects. The locally affine invariance is achieved by simple linear algebra for efficient and robust computations, making the algorithm suitable for detections under object distortions like perspective projection. Commonly used edge detectors and a voting process are, respectively, used before and after the edge description and matching steps to form a complete reflection detection pipeline. Experiments are performed using synthetic and real-world images with both multiple and single reflection symmetry axis. The test results are compared with existing algorithms to validate the proposed method. PMID:25608306

  18. Effects of whole-body irradiation on antibody affinity.

    PubMed Central

    Gorini, G; Adorini, L; Boraschi, D; Di Michele, A; Doria, G

    1977-01-01

    Mice exposed to a sublethal dose of X-rays were immunized with alum-precipitated DNP-KLH (dinitrophenyl-keyhole limpet haemocyanin) and B. pertussis either before or after irradiation. The primary anti-DNP antibody response was evaluated during 8 weeks after immunization by the equilibrium dialysis technique using ammonium sulphate- precipitated serum globulins and the ligand 3H-labelled xi-DNP-L-Lysine. The serum concentrations of antibody sites in mice immunized 1-5 days before or 2 h-8 weeks after 450 rad were below the values in unirradiated controls at all bleeding times. Antibody affinity, however, was found to be up to 20 fold higher in irradiated mice than in control mice when antigen was injected before, or 3-8 weeks after, irradiation. Spleen cells from mice exposed to 450 rad 1-9 weeks before killing were stimulated in vitro with PHA, ConA, or LPS. Recovery profiles of mitotic responsiveness suggest that enhancement of antibody affinity in irradiated mice could result from relative lack of suppressor T Cells. PMID:198358

  19. Affinity purification of copper chelating peptides from chickpea protein hydrolysates.

    PubMed

    Megías, Cristina; Pedroche, Justo; Yust, Maria M; Girón-Calle, Julio; Alaiz, Manuel; Millan, Francisco; Vioque, Javier

    2007-05-16

    Chickpea protein hydrolysates obtained with alcalase and flavourzyme were used for purification of copper chelating peptides by affinity chromatography using copper immobilized on solid supports. The chelating activity of purified peptides was indirectly measured by the inhibition of beta-carotene oxidation in the presence of copper. Two protein hydrolysates, obtained after 10 and 100 min of hydrolysis, were the most inhibitory of beta-carotene oxidation. Purified copper chelating peptides from these protein hydrolysates contained 19.7 and 35.1% histidine, respectively, in comparison to 2.7 and 2.6% in the protein hydrolysates. Chelating peptides from hydrolysate obtained after 10 min of hydrolysis were the most antioxidative being 8.3 times more antioxidative than the hydrolysate, while chelating peptides purified from protein hydrolysate obtained after 100 min were 3.1 times more antioxidative than its hydrolysate. However, the histidine content was higher in peptides derived from the 100 min hydrolysate (19.7 against 35.1% in 10 min hydrolysate), indicating that this amino acid is not the only factor involved in the antioxidative activity, and other factors such as peptide size or amino acid sequence are also determinant. This manuscript shows that affinity chromatography is a useful procedure for purification of copper chelating peptides. This method can be extended to other metals of interest in nutrition, such as calcium, iron, or zinc. Purified chelating peptides, in addition to their antioxidative properties, may also be useful in food mineral fortification for increasing the bioavailability of these metals.

  20. Compensating Enthalpic and Entropic Changes Hinder Binding Affinity Optimization

    SciTech Connect

    Lafont,V.; Armstrong, A.; Ohtaka, H.; Kiso, Y.; Amzel, L.; Freire, E.

    2007-01-01

    A common strategy to improve the potency of drug candidates is to introduce chemical functionalities, like hydrogen bond donors or acceptors, at positions where they are able to establish strong interactions with the target. However, it is often observed that the added functionalities do not necessarily improve potency even if they form strong hydrogen bonds. Here, we explore the thermodynamic and structural basis for those observations. KNI-10033 is a potent experimental HIV-1 protease inhibitor with picomolar affinity against the wild-type enzyme (Kd = 13 pm). The potency of the inhibitor is the result of favorable enthalpic (?H = -8.2 kcal/mol) and entropic (-T?S = -6.7 kcal/mol) interactions. The replacement of the thioether group in KNI-10033 by a sulfonyl group (KNI-10075) results in a strong hydrogen bond with the amide of Asp 30B of the HIV-1 protease. This additional hydrogen bond improves the binding enthalpy by 3.9 kcal/mol; however, the enthalpy gain is completely compensated by an entropy loss, resulting in no affinity change. Crystallographic and thermodynamic analysis of the inhibitor/protease complexes indicates that the entropy losses are due to a combination of conformational and solvation effects. These results provide a set of practical guidelines aimed at overcoming enthalpy/entropy compensation and improve binding potency.

  1. Benzodiazepines: electron affinity, receptors and cell signaling - a multifaceted approach.

    PubMed

    Kovacic, Peter; Ott, Nadia; Cooksy, Andrew L

    2013-12-01

    This report entails a multifaceted approach to benzodiazepine (BZ) action, involving electron affinity, receptors, cell signaling and other aspects. Computations of the electron affinities (EAs) of different BZs have been carried out to establish the effect of various substituents on their EA. These computations were undertaken to serve as a first step in determining what role electron transfer (ET) plays in BZ activity. The calculations were conducted on the premise that the nature of the substituent will either decrease or increase the electron density of the benzene ring, thus altering the ability of the molecule to accept an electron. Investigations were performed on the effect of drug protonation on EA. Similarities involving substituent effects in prior electrochemical studies are also discussed. As part of the multifaceted approach, EA is linked to ET, which appears to play a role in therapeutic activity and toxicity. There is extensive literature dealing with the role of receptors in BZ activity. Significant information on receptor involvement was reported more than 40 years ago. Gamma-aminobutyric acid (GABA) is known to be importantly involved. GABA is a probable mediator of BZ effects. BZ and GABA receptors, although not identical, are physiologically linked. Cell signaling is known to play a part in the biochemistry of BZ action. Various factors participated, such as gene expression, allosteric influence, toxic effects and therapeutic action. Evidence points to involvement of EA and ET in the mode of action in cell signaling. Oxidative stress and antioxidant effects are also addressed.

  2. Robust activation method for negative electron affinity photocathodes

    DOEpatents

    Mulhollan, Gregory A.; Bierman, John C.

    2011-09-13

    A method by which photocathodes(201), single crystal, amorphous, or otherwise ordered, can be surface modified to a robust state of lowered and in best cases negative, electron affinity has been discovered. Conventional methods employ the use of Cs(203) and an oxidizing agent(207), typically carried by diatomic oxygen or by more complex molecules, for example nitrogen trifluoride, to achieve a lowered electron affinity(404). In the improved activation method, a second alkali, other than Cs(205), is introduced onto the surface during the activation process, either by co-deposition, yo-yo, or sporadic or intermittent application. Best effect for GaAs photocathodes has been found through the use of Li(402) as the second alkali, though nearly the same effect can be found by employing Na(406). Suitable photocathodes are those which are grown, cut from boules, implanted, rolled, deposited or otherwise fabricated in a fashion and shape desired for test or manufacture independently supported or atop a support structure or within a framework or otherwise affixed or suspended in the place and position required for use.

  3. Flexible Molybdenum Electrodes towards Designing Affinity Based Protein Biosensors.

    PubMed

    Kamakoti, Vikramshankar; Panneer Selvam, Anjan; Radha Shanmugam, Nandhinee; Muthukumar, Sriram; Prasad, Shalini

    2016-01-01

    Molybdenum electrode based flexible biosensor on porous polyamide substrates has been fabricated and tested for its functionality as a protein affinity based biosensor. The biosensor performance was evaluated using a key cardiac biomarker; cardiac Troponin-I (cTnI). Molybdenum is a transition metal and demonstrates electrochemical behavior upon interaction with an electrolyte. We have leveraged this property of molybdenum for designing an affinity based biosensor using electrochemical impedance spectroscopy. We have evaluated the feasibility of detection of cTnI in phosphate-buffered saline (PBS) and human serum (HS) by measuring impedance changes over a frequency window from 100 mHz to 1 MHz. Increasing changes to the measured impedance was correlated to the increased dose of cTnI molecules binding to the cTnI antibody functionalized molybdenum surface. We achieved cTnI detection limit of 10 pg/mL in PBS and 1 ng/mL in HS medium. The use of flexible substrates for designing the biosensor demonstrates promise for integration with a large-scale batch manufacturing process. PMID:27438863

  4. Boronate affinity adsorption of RNA: possible role of conformational changes

    NASA Technical Reports Server (NTRS)

    Singh, N.; Willson, R. C.; Fox, G. E. (Principal Investigator)

    1999-01-01

    Batch equilibrium adsorption isotherm determination is used to characterize the adsorption of mixed yeast RNA on agarose-immobilized m-aminophenylboronic acid. It is shown that the affinity-enhancing influence of divalent cations depends strongly on the precise nature of the cation used, with barium being far more effective than the conventionally-used magnesium. This adsorption-promoting influence of barium is suggested to arise primarily from ionic influences on the structure and rigidity of the RNA molecule, as the adsorption of ribose-based small molecules is not similarly affected. The substitution of barium for the standard magnesium counterion does not greatly promote the adsorption of DNA, implying that the effect is specific to RNA and may be useful in boronate-based RNA separations. RNA adsorption isotherms exhibit sharp transitions as functions of temperature, and these transitions occur at different temperatures with Mg2+ and with Ba2+. Adsorption affinity and capacity were found to increase markedly at lower temperatures, suggestive of an enthalpically favored interaction process. The stoichiometric displacement parameter, Z, in Ba2+ buffer is three times the value in Mg2+ buffer, and is close to unity.

  5. 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).

  6. Flexible Molybdenum Electrodes towards Designing Affinity Based Protein Biosensors

    PubMed Central

    Kamakoti, Vikramshankar; Panneer Selvam, Anjan; Radha Shanmugam, Nandhinee; Muthukumar, Sriram; Prasad, Shalini

    2016-01-01

    Molybdenum electrode based flexible biosensor on porous polyamide substrates has been fabricated and tested for its functionality as a protein affinity based biosensor. The biosensor performance was evaluated using a key cardiac biomarker; cardiac Troponin-I (cTnI). Molybdenum is a transition metal and demonstrates electrochemical behavior upon interaction with an electrolyte. We have leveraged this property of molybdenum for designing an affinity based biosensor using electrochemical impedance spectroscopy. We have evaluated the feasibility of detection of cTnI in phosphate-buffered saline (PBS) and human serum (HS) by measuring impedance changes over a frequency window from 100 mHz to 1 MHz. Increasing changes to the measured impedance was correlated to the increased dose of cTnI molecules binding to the cTnI antibody functionalized molybdenum surface. We achieved cTnI detection limit of 10 pg/mL in PBS and 1 ng/mL in HS medium. The use of flexible substrates for designing the biosensor demonstrates promise for integration with a large-scale batch manufacturing process. PMID:27438863

  7. Binding Affinity of Glycoconjugates to BACILLUS Spores and Toxins

    NASA Astrophysics Data System (ADS)

    Rasol, Aveen; Eassa, Souzan; Tarasenko, Olga

    2010-04-01

    Early recognition of Bacillus cereus group species is important since they can cause food-borne illnesses and deadly diseases in humans. Glycoconjugates (GCs) are carbohydrates covalently linked to non-sugar moieties including lipids, proteins or other entities. GCs are involved in recognition and signaling processes intrinsic to biochemical functions in cells. They also stimulate cell-cell adhesion and subsequent recognition and activation of receptors. We have demonstrated that GCs are involved in Bacillus cereus spore recognition. In the present study, we have investigated whether GCs possess the ability to bind and recognize B. cereus spores and Bacillus anthracis recombinant single toxins (sTX) and complex toxins (cTX). The affinity of GCs to spores + sTX and spores + cTX toxins was studied in the binding essay. Our results demonstrated that GC9 and GC10 were able to selectively bind to B. cereus spores and B. anthracis toxins. Different binding affinities for GCs were found toward Bacillus cereus spores + sTX and spores + cTX. Dilution of GCs does not impede the recognition and binding. Developed method provides a tool for simultaneous recognition and targeting of spores, bacteria toxins, and/or other entities.

  8. Affinity labeling of GTP-binding proteins in cellular extracts.

    PubMed

    Löw, A; Faulhammer, H G; Sprinzl, M

    1992-05-25

    GTP-binding proteins in cellular extracts from Escherichia coli, Thermus thermophilus, yeast, wheat germ or calf thymus were identified using in situ periodate-oxidized [alpha-32P]GTP as affinity label. Site-specific reaction of individual GTP-binding proteins was achieved by cross-linking the protein-bound 2',3'-dialdehyde derivative of GTP with the single lysine residue of the conserved NKXD sequence through Schiff's base formation and subsequent cyanoborohydride reduction. Labeled GTP-binding proteins from prokaryotic or eukaryotic cell homogenates were separated by polyacrylamide gel electrophoresis and visualized by autoradiography. In addition cross-linking of [alpha-32P]GTP with GTP-binding proteins was demonstrated in model systems using different purified GTPases, human c-H-ras p21, transducin from bovine retina, polypeptide elongation factor Tu (EF-Tu) from T. thermophilus and initiation factor 2 (IF2) from T. thermophilus. The described affinity labeling technique can serve as an analytical method for the identification of GTPases belonging to the classes of ras-proteins, elongation and initiation factors, and heterotrimeric signal transducing G-proteins. PMID:1592117

  9. Telonemia, a new protist phylum with affinity to chromist lineages.

    PubMed

    Shalchian-Tabrizi, K; Eikrem, W; Klaveness, D; Vaulot, D; Minge, M A; Le Gall, F; Romari, K; Throndsen, J; Botnen, A; Massana, R; Thomsen, H A; Jakobsen, K S

    2006-07-22

    Recent molecular investigations of marine samples taken from different environments, including tropical, temperate and polar areas, as well as deep thermal vents, have revealed an unexpectedly high diversity of protists, some of them forming deep-branching clades within important lineages, such as the alveolates and heterokonts. Using the same approach on coastal samples, we have identified a novel group of protist small subunit (SSU) rDNA sequences that do not correspond to any phylogenetic group previously identified. Comparison with other sequences obtained from cultures of heterotrophic protists showed that the environmental sequences grouped together with Telonema, a genus known since 1913 but of uncertain taxonomic affinity. Phylogenetic analyses using four genes (SSU, Hsp90, alpha-tubulin and beta-tubulin), and accounting for gamma- and covarion-distributed substitution rates, revealed Telonema as a distinct group of species branching off close to chromist lineages. Consistent with these gene trees, Telonema possesses ultrastructures revealing both the distinctness of the group and the evolutionary affinity to chromist groups. Altogether, the data suggest that Telonema constitutes a new eukaryotic phylum, here defined as Telonemia, possibly representing a key clade for the understanding of the early evolution of bikont protist groups, such as the proposed chromalveolate supergroup.

  10. Altered catecholamine receptor affinity in rabbit aortic intimal hyperplasia

    SciTech Connect

    O'Malley, M.K.; Cotecchia, S.; Hagen, P.O. )

    1991-08-01

    Intimal thickening is a universal response to endothelial denudation and is also thought to be a precursor of atherosclerosis. The authors have demonstrated selective supersensitivity in arterial intimal hyperplasia to norepinephrine and they now report a possible mechanism for this. Binding studies in rabbit aorta with the selective alpha 1-adrenergic radioligand 125I-HEAT demonstrated that there was no change in receptor density (20 {plus minus} 4 fmole/10(6) cells) in intact vascular smooth muscle cells at either 5 or 14 days after denudation. However, competition studies showed a 2.6-fold increase in alpha 1-adrenergic receptor affinity for norepinephrine in intimal hyperplastic tissue (P less than 0.05). This increased affinity for norepinephrine was associated with a greater increase in 32P-labeled phosphatidylinositol (148% intimal thickening versus 76% control) and phosphatidic acid (151% intimal thickening versus 56% control) following norepinephrine stimulation of free floating rings of intimal hyperplastic aorta. These data suggest that the catecholamine supersensitivity in rabbit aortic intimal hyperplasia is receptor mediated and may be linked to the phosphatidylinositol cycle.

  11. A global benchmark study using affinity-based biosensors.

    PubMed

    Rich, Rebecca L; Papalia, Giuseppe A; Flynn, Peter J; Furneisen, Jamie; Quinn, John; Klein, Joshua S; Katsamba, Phini S; Waddell, M Brent; Scott, Michael; Thompson, Joshua; Berlier, Judie; Corry, Schuyler; Baltzinger, Mireille; Zeder-Lutz, Gabrielle; Schoenemann, Andreas; Clabbers, Anca; Wieckowski, Sebastien; Murphy, Mary M; Page, Phillip; Ryan, Thomas E; Duffner, Jay; Ganguly, Tanmoy; Corbin, John; Gautam, Satyen; Anderluh, Gregor; Bavdek, Andrej; Reichmann, Dana; Yadav, Satya P; Hommema, Eric; Pol, Ewa; Drake, Andrew; Klakamp, Scott; Chapman, Trevor; Kernaghan, Dawn; Miller, Ken; Schuman, Jason; Lindquist, Kevin; Herlihy, Kara; Murphy, Michael B; Bohnsack, Richard; Andrien, Bruce; Brandani, Pietro; Terwey, Danny; Millican, Rohn; Darling, Ryan J; Wang, Liann; Carter, Quincy; Dotzlaf, Joe; Lopez-Sagaseta, Jacinto; Campbell, Islay; Torreri, Paola; Hoos, Sylviane; England, Patrick; Liu, Yang; Abdiche, Yasmina; Malashock, Daniel; Pinkerton, Alanna; Wong, Melanie; Lafer, Eileen; Hinck, Cynthia; Thompson, Kevin; Primo, Carmelo Di; Joyce, Alison; Brooks, Jonathan; Torta, Federico; Bagge Hagel, Anne Birgitte; Krarup, Janus; Pass, Jesper; Ferreira, Monica; Shikov, Sergei; Mikolajczyk, Malgorzata; Abe, Yuki; Barbato, Gaetano; Giannetti, Anthony M; Krishnamoorthy, Ganeshram; Beusink, Bianca; Satpaev, Daulet; Tsang, Tiffany; Fang, Eric; Partridge, James; Brohawn, Stephen; Horn, James; Pritsch, Otto; Obal, Gonzalo; Nilapwar, Sanjay; Busby, Ben; Gutierrez-Sanchez, Gerardo; Gupta, Ruchira Das; Canepa, Sylvie; Witte, Krista; Nikolovska-Coleska, Zaneta; Cho, Yun Hee; D'Agata, Roberta; Schlick, Kristian; Calvert, Rosy; Munoz, Eva M; Hernaiz, Maria Jose; Bravman, Tsafir; Dines, Monica; Yang, Min-Hsiang; Puskas, Agnes; Boni, Erica; Li, Jiejin; Wear, Martin; Grinberg, Asya; Baardsnes, Jason; Dolezal, Olan; Gainey, Melicia; Anderson, Henrik; Peng, Jinlin; Lewis, Mark; Spies, Peter; Trinh, Quyhn; Bibikov, Sergei; Raymond, Jill; Yousef, Mohammed; Chandrasekaran, Vidya; Feng, Yuguo; Emerick, Anne; Mundodo, Suparna; Guimaraes, Rejane; McGirr, Katy; Li, Yue-Ji; Hughes, Heather; Mantz, Hubert; Skrabana, Rostislav; Witmer, Mark; Ballard, Joshua; Martin, Loic; Skladal, Petr; Korza, George; Laird-Offringa, Ite; Lee, Charlene S; Khadir, Abdelkrim; Podlaski, Frank; Neuner, Phillippe; Rothacker, Julie; Rafique, Ashique; Dankbar, Nico; Kainz, Peter; Gedig, Erk; Vuyisich, Momchilo; Boozer, Christina; Ly, Nguyen; Toews, Mark; Uren, Aykut; Kalyuzhniy, Oleksandr; Lewis, Kenneth; Chomey, Eugene; Pak, Brian J; Myszka, David G

    2009-03-15

    To explore the variability in biosensor studies, 150 participants from 20 countries were given the same protein samples and asked to determine kinetic rate constants for the interaction. We chose a protein system that was amenable to analysis using different biosensor platforms as well as by users of different expertise levels. The two proteins (a 50-kDa Fab and a 60-kDa glutathione S-transferase [GST] antigen) form a relatively high-affinity complex, so participants needed to optimize several experimental parameters, including ligand immobilization and regeneration conditions as well as analyte concentrations and injection/dissociation times. Although most participants collected binding responses that could be fit to yield kinetic parameters, the quality of a few data sets could have been improved by optimizing the assay design. Once these outliers were removed, the average reported affinity across the remaining panel of participants was 620 pM with a standard deviation of 980 pM. These results demonstrate that when this biosensor assay was designed and executed appropriately, the reported rate constants were consistent, and independent of which protein was immobilized and which biosensor was used.

  12. Characterization of methacrylate chromatographic monoliths bearing affinity ligands.

    PubMed

    Černigoj, Urh; Vidic, Urška; Nemec, Blaž; Gašperšič, Jernej; Vidič, Jana; Lendero Krajnc, Nika; Štrancar, Aleš; Podgornik, Aleš

    2016-09-16

    We investigated effect of immobilization procedure and monolith structure on chromatographic performance of methacrylate monoliths bearing affinity ligands. Monoliths of different pore size and various affinity ligands were prepared and characterized using physical and chromatographic methods. When testing protein A monoliths with different protein A ligand densities, a significant nonlinear effect of ligand density on dynamic binding capacity (DBC) for IgG was obtained and accurately described by Langmuir isotherm curve enabling estimation of protein A utilization as a function of ligand density. Maximal IgG binding capacity was found to be at least 12mg/mL exceeding theoretical monolayer adsorption value of 7.8mg/mL assuming hexagonal packing and IgG hydrodynamic diameter of 11nm. Observed discrepancy was explained by shrinkage of IgG during adsorption on protein A experimentally determined through calculated adsorbed IgG layer thickness of 5.4nm from pressure drop data. For monoliths with different pore size maximal immobilized densities of protein A as well as IgG dynamic capacity linearly correlates with monolith surface area indicating constant ligand utilization. Finally, IgGs toward different plasma proteins were immobilized via the hydrazide coupling chemistry to provide oriented immobilization. DBC was found to be flow independent and was increasing with the size of bound protein. Despite DBC was lower than IgG capacity to immobilized protein A, ligand utilization was higher. PMID:27554023

  13. 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.

  14. Membrane Affinity of Platensimycin and Its Dialkylamine Analogs

    PubMed Central

    Rowe, Ian; Guo, Min; Yasmann, Anthony; Cember, Abigail; Sintim, Herman O.; Sukharev, Sergei

    2015-01-01

    Membrane permeability is a desired property in drug design, but there have been difficulties in quantifying the direct drug partitioning into native membranes. Platensimycin (PL) is a new promising antibiotic whose biosynthetic production is costly. Six dialkylamine analogs of PL were synthesized with identical pharmacophores but different side chains; five of them were found inactive. To address the possibility that their activity is limited by the permeation step, we calculated polarity, measured surface activity and the ability to insert into the phospholipid monolayers. The partitioning of PL and the analogs into the cytoplasmic membrane of E. coli was assessed by activation curve shifts of a re-engineered mechanosensitive channel, MscS, in patch-clamp experiments. Despite predicted differences in polarity, the affinities to lipid monolayers and native membranes were comparable for most of the analogs. For PL and the di-myrtenyl analog QD-11, both carrying bulky sidechains, the affinity for the native membrane was lower than for monolayers (half-membranes), signifying that intercalation must overcome the lateral pressure of the bilayer. We conclude that the biological activity among the studied PL analogs is unlikely to be limited by their membrane permeability. We also discuss the capacity of endogenous tension-activated channels to detect asymmetric partitioning of exogenous substances into the native bacterial membrane and the different contributions to the thermodynamic force which drives permeation. PMID:26247942

  15. The eyes of Tullimonstrum reveal a vertebrate affinity.

    PubMed

    Clements, Thomas; Dolocan, Andrei; Martin, Peter; Purnell, Mark A; Vinther, Jakob; Gabbott, Sarah E

    2016-04-28

    Tullimonstrum gregarium is an iconic soft-bodied fossil from the Carboniferous Mazon Creek Lagerstätte (Illinois, USA). Despite a large number of specimens and distinct anatomy, various analyses over the past five decades have failed to determine the phylogenetic affinities of the 'Tully monster', and although it has been allied to such disparate phyla as the Mollusca, Annelida or Chordata, it remains enigmatic. The nature and phylogenetic affinities of Tullimonstrum have defied confident systematic placement because none of its preserved anatomy provides unequivocal evidence of homology, without which comparative analysis fails. Here we show that the eyes of Tullimonstrum possess ultrastructural details indicating homology with vertebrate eyes. Anatomical analysis using scanning electron microscopy reveals that the eyes of Tullimonstrum preserve a retina defined by a thick sheet comprising distinct layers of spheroidal and cylindrical melanosomes. Time-of-flight secondary ion mass spectrometry and multivariate statistics provide further evidence that these microbodies are melanosomes. A range of animals have melanin in their eyes, but the possession of melanosomes of two distinct morphologies arranged in layers, forming retinal pigment epithelium, is a synapomorphy of vertebrates. Our analysis indicates that in addition to evidence of colour patterning, ecology and thermoregulation, fossil melanosomes can also carry a phylogenetic signal. Identification in Tullimonstrum of spheroidal and cylindrical melanosomes forming the remains of retinal pigment epithelium indicates that it is a vertebrate; considering its body parts in this new light suggests it was an anatomically unusual member of total group Vertebrata. PMID:27074512

  16. Affinity-based screening techniques for enhancing lead discovery.

    PubMed

    Comess, Kenneth M; Schurdak, Mark E

    2004-07-01

    Contemporary, rational small-molecule lead discovery methods, comprising target identification, assay development, high-throughput screening (HTS), hit characterization and medicinal chemistry optimization, dominate early-stage drug discovery strategies in many pharmaceutical companies. There is a growing disparity between the increasing cost of funding these methods and the decreasing number of new drugs reaching the market. New strategies must be adopted to reverse this trend. The use of genomics- and proteomics-based target discovery efforts can aid the process by dramatically increasing the number of novel, more highly validated targets entering the discovery process, but HTS must meet this increased demand with faster, cheaper technologies. Although activity-based screening strategies are typically efficient, allowing one scientist to interrogate tens of thousands of compounds per day, affinity-based screening strategies can allow much greater efficiency in the overall process. Affinity-based methods can play a role in both facilitating the screening of a greater number of targets and in efficiently characterizing the primary hits discovered.

  17. Cholinergic activation of the murine trachealis muscle via non-vesicular acetylcholine release involving low-affinity choline transporters.

    PubMed

    Nassenstein, Christina; Wiegand, Silke; Lips, Katrin S; Li, Guanfeng; Klein, Jochen; Kummer, Wolfgang

    2015-11-01

    In addition to quantal, vesicular release of acetylcholine (ACh), there is also non-quantal release at the motor endplate which is insufficient to evoke postsynaptic responses unless acetylcholinesterase (AChE) is inhibited. We here addressed potential non-quantal release in the mouse trachea by organ bath experiments and (immuno)histochemical methods. Electrical field stimulation (EFS) of nerve terminals elicited tracheal constriction that is largely due to ACh release. Classical enzyme histochemistry demonstrated acetylcholinesterase (AChE) activity in nerve fibers in the muscle and butyrylcholinesterase (BChE) activity in the smooth muscle cells. Acute inhibition of both esterases by eserine significantly raised tracheal tone which was fully sensitive to atropine. This effect was reduced, but not abolished, in AChE, but not in BChE gene-deficient mice. The eserine-induced increase in tracheal tone was unaffected by vesamicol (10(-5)M), an inhibitor of the vesicular acetylcholine transporter, and by corticosterone (10(-4)M), an inhibitor of organic cation transporters. Hemicholinium-3, in low concentrations an inhibitor of the high-affinity choline transporter-1 (CHT1), completely abrogated the eserine effects when applied in high concentrations (10(-4)M) pointing towards an involvement of low-affinity choline transporters. To evaluate the cellular sources of non-quantal ACh release in the trachea, expression of low-affinity choline transporter-like family (CTL1-5) was evaluated by RT-PCR analysis. Even though these transporters were largely abundant in the epithelium, denudation of airway epithelial cells had no effect on eserine-induced tracheal contraction, indicating a non-quantal release of ACh from non-epithelial sources in the airways. These data provide evidence for an epithelium-independent non-vesicular, non-quantal ACh release in the mouse trachea involving low-affinity choline transporters. PMID:26278668

  18. 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

  19. Thermodynamic mixing of molecular states of the epidermal growth factor receptor modulates macroscopic ligand binding affinity.

    PubMed Central

    Holbrook, M R; Slakey, L L; Gross, D J

    2000-01-01

    The epidermal growth factor receptor (EGFr), when expressed on the cell surface, has long been known to display two distinct affinities for epidermal growth factor (EGF) binding. In addition, the treatment of cells expressing the EGFr with phorbol esters has been shown to cause a loss of the high-affinity binding capacity of the receptor. In the present study, point mutations that alter acidic or phosphorylation sites have been made in an intracellular domain near Tyr-992 (residues 988-992) of the EGFr. Equilibrium (125)I-EGF binding studies demonstrate that the conversion of Tyr-992 into glutamate induces a 4-fold decrease in the EGFr apparent low-affinity dissociation constant, whereas the mutation of two acidic residues, Asp-988 and Glu-991, or the conversion of Tyr-992 into phenylalanine does not alter EGFr affinity. Phorbol ester treatment of EGFr-expressing Chinese hamster ovary cells results in a loss of high-affinity binding and an increase in the apparent low-affinity dissociation constant of the receptor, similar to the effect of a truncation mutant in which the C-terminal 190 residues are deleted. These results are examined in the context of a new model for regulation of the affinity of the EGFr for EGF in which a cytosolic particle stabilizes the high-affinity conformation of the EGFr and a rapid equilibrium exists between EGFr high-affinity and low-affinity conformations. This model demonstrates that the macroscopic affinities of the EGFr can differ from the affinities of individual EGFr molecules and provides a theoretical framework whereby the measured affinities of the EGFr are modulated by intracellular interactions. PMID:11062062

  20. On a solitonic specialisation for the general solutions of some two-dimensional completely integrable systems

    NASA Astrophysics Data System (ADS)

    I. Olive, David; V. Saveliev, Mikhail; W. R. Underwood, Jonathan

    1993-07-01

    Following a prescription of Olive, Turok and Underwood for a solitonic specialisation of the general solutions to the (abelian) periodic Toda field theories, we discuss a construction of the soliton solutions for a wide class of two-dimensional completely integrable systems arising in the framework of the group-algebraic approach, including the “non-abelian” version of the affine Toda theory.

  1. Chelators whose affinity for calcium is decreased by illumination

    NASA Technical Reports Server (NTRS)

    Tsien, Roger Y. (Inventor); Grynkiewicz, Grzegorz (Inventor); Minta, Akwasi (Inventor)

    1987-01-01

    The present invention discloses a group of calcium chelating compounds which have a descreased affinity for calcium following illumination. These new compounds contain a photolabile nitrobenzyl derivative coupled to a tetracarboxylate Ca.sup.2+ chelating parent compound having the octacoordinate chelating groups characteristic of EGTA or BAPTA. In a first form, the new compounds are comprised of a BAPTA-like chelator coupled to a single 2-nitrobenzyl derivative, which in turn is a photochemical precursor of a 2-nitrosobenzophenone. In a second form, the new compounds are comprised of a BAPTA-like chelator coupled to two 2-nitrobenzyl derivatives, themselves photochemical prcursors of the related 2-nitrosobenzophenones. The present invention also discloses a novel method for preparing 1-hydroxy- or 1-alkoxy-1-(2-nitroaryl)-1-aryl methanes. Methanes of this type are critical to the preparation of, or actually constitute, the photolabile Ca.sup.2+ chelating compounds disclosed and claimed herein.

  2. Deformation of supersymmetric and conformal quantum mechanics through affine transformations

    NASA Astrophysics Data System (ADS)

    Spiridonov, Vyacheslav

    Affine transformations (dilatations and translations) are used to define a deformation of one-dimensional N = 2 supersymmetric quantum mechanics. Resulting physical systems do not have conserved charges and degeneracies in the spectra. Instead, superpartner Hamiltonians are q-isospectral, i.e. the spectrum of one can be obtained from another (with possible exception of the lowest level) by q(sup 2)-factor scaling. This construction allows easily to rederive a special self-similar potential found by Shabat and to show that for the latter a q-deformed harmonic oscillator algebra of Biedenharn and Macfarlane serves as the spectrum generating algebra. A general class of potentials related to the quantum conformal algebra su(sub q)(1,1) is described. Further possibilities for q-deformation of known solvable potentials are outlined.

  3. Growth factors with heparin binding affinity in human synovial fluid

    SciTech Connect

    Hamerman, D.; Taylor, S.; Kirschenbaum, I.; Klagsbrun, M.; Raines, E.W.; Ross, R.; Thomas, K.A.

    1987-12-01

    Synovial effusions were obtained from the knees of 15 subjects with joint trauma, menisceal or ligamentous injury, or osteoarthritis. Heparin-Sepharose affinity chromatography of these synovial fluids revealed, in general, three major peaks of mitogenic activity as measured by incorporation of /sup 3/H-thymidine into 3T3 cells. Gradient elution patterns showed activities at 0.5M NaCl, which is characteristic of platelet derived growth factor, and at 1.1 M NaCl and 1.6M NaCl, indicative of acidic and basic fibroblast growth factors, respectively. The identities of these mitogenic fractions were confirmed by specific immunologic and receptor-binding assays. The presence of platelet derived, acidic and basic fibroblast growth factors in the synovial fluid may contribute to wound healing in the arthritic joint.

  4. Robust template matching for affine resistant image watermarks.

    PubMed

    Pereira, S; Pun, T

    2000-01-01

    Digital watermarks have been proposed as a method for discouraging illicit copying and distribution of copyrighted material. This paper describes a method for the secure and robust copyright protection of digital images. We present an approach for embedding a digital watermark into an image using the Fourier transform. To this watermark is added a template in the Fourier transform domain to render the method robust against general linear transformations. We detail a new algorithm based on polar maps for the accurate and efficient recovery of the template in an image which has undergone a general affine transformation. We also present results which demonstrate the robustness of the method against some common image processing operations such as compression, rotation, scaling, and aspect ratio changes. PMID:18255481

  5. Identification of protein interacting partners using tandem affinity purification.

    PubMed

    Bailey, Dalan; Urena, Luis; Thorne, Lucy; Goodfellow, Ian

    2012-01-01

    A critical and often limiting step in understanding the function of host and viral proteins is the identification of interacting cellular or viral protein partners. There are many approaches that allow the identification of interacting partners, including the yeast two hybrid system, as well as pull down assays using recombinant proteins and immunoprecipitation of endogenous proteins followed by mass spectrometry identification(1). Recent studies have highlighted the utility of double-affinity tag mediated purification, coupled with two specific elution steps in the identification of interacting proteins. This approach, termed Tandem Affinity Purification (TAP), was initially used in yeast(2,3) but more recently has been adapted to use in mammalian cells(4-8). As proof-of-concept we have established a tandem affinity purification (TAP) method using the well-characterized eukaryotic translation initiation factor eIF4E(9,10).The cellular translation factor eIF4E is a critical component of the cellular eIF4F complex involved in cap-dependent translation initiation(10). The TAP tag used in the current study is composed of two Protein G units and a streptavidin binding peptide separated by a Tobacco Etch Virus (TEV) protease cleavage sequence. The TAP tag used in the current study is composed of two Protein G units and a streptavidin binding peptide separated by a Tobacco Etch Virus (TEV) protease cleavage sequence(8). To forgo the need for the generation of clonal cell lines, we developed a rapid system that relies on the expression of the TAP-tagged bait protein from an episomally maintained plasmid based on pMEP4 (Invitrogen). Expression of tagged murine eIF4E from this plasmid was controlled using the cadmium chloride inducible metallothionein promoter. Lysis of the expressing cells and subsequent affinity purification via binding to rabbit IgG agarose, TEV protease cleavage, binding to streptavidin linked agarose and subsequent biotin elution identified numerous

  6. Intracellular calcium ions decrease the affinity of the GABA receptor.

    PubMed

    Inoue, M; Oomura, Y; Yakushiji, T; Akaike, N

    Intracellular free Ca2+ [( Ca2+]i) plays a crucial role in the transduction of extracellular signals. It has been implicated in the modulation of light sensitivity in Limulus photoreceptors and in the efficacy of synaptic transmission; calcium ion fluxes are also involved in the postsynaptic facilitation of nicotinic transmission seen in sympathetic ganglia, and in activation of the acetylcholine (ACh) receptor. [Ca2+]i is also a second messenger for many biologically active substances. We recorded neuronal activities of sensory neurones from the bullfrog (Rana catesbiana), using the suction pipette method and a 'concentration clamp' technique to apply gamma-aminobutyric acid (GABA) to the cell. We report the first evidence that [Ca2+]i suppresses the GABA-activated Cl- conductance, by decreasing the apparent affinity of the GABA receptor. PMID:2431316

  7. Approximated affine projection algorithm for feedback cancellation in hearing aids.

    PubMed

    Lee, Sangmin; Kim, In-Young; Park, Young-Cheol

    2007-09-01

    We propose an approximated affine projection (AP) algorithm for feedback cancellation in hearing aids. It is based on the conventional approach using the Gauss-Seidel (GS) iteration, but provides more stable convergence behaviour even with small step sizes. In the proposed algorithm, a residue of the weighted error vector, instead of the current error sample, is used to provide stable convergence. A new learning rate control scheme is also applied to the proposed algorithm to prevent signal cancellation and system instability. The new scheme determines step size in proportion to the prediction factor of the input, so that adaptation is inhibited whenever tone-like signals are present in the input. Simulation results verified the efficiency of the proposed algorithm. PMID:17644214

  8. Bioadhesive nanoareas in antifouling matrix for highly efficient affinity sensors

    NASA Astrophysics Data System (ADS)

    Mannelli, I.; Valsesia, A.; Lisboa, P.; Anger-Leroy, M.; Mornet, S.; Colpo, P.; Rossi, F.

    2008-08-01

    A procedure for fabricating nanopatterned surfaces at the sub-500 nm scale comprising a hexagonal close packed array of bioadhesive gold nanoareas in a protein resistant matrix (PEO-like polymer), has been optimized. The surfaces were characterized by AFM analysis and their interaction with amino functionalised gold nanoparticles as models were investigated. The AFM images show the crystalline arrangement of nanopattern array and the localized adsorption of the H2N-Au nanoparticles in the bioadhesive nanoareas. A Surface Plasmon Resonance imaging (SPRi) system was used to assess the detection performances of these surfaces when employed as a transduction platform for studying biomolecule interactions. The investigated surfaces showed an enhancement of the affinity reaction efficiency with respect to the non structured surfaces. The obtained preliminary results show that nanostructuring the surfaces improve the binding site accessibility of the immobilized biological probes without significantly modifying the native biomolecule conformation.

  9. Bioavailable affinity label for collagen prolyl 4-hydroxylase

    PubMed Central

    Vasta, James D.; Higgin, Joshua J.; Kersteen, Elizabeth A.

    2013-01-01

    Collagen is the most abundant protein in animals. Its prevalent 4-hydroxyproline residues contribute greatly to its conformational stability. The hydroxyl groups arise from a post-translational modification catalyzed by the non-heme iron-dependent enzyme, collagen prolyl 4-hydroxylase (P4H). Here, we report that 4-oxo-5,6-epoxyhexanoate, a mimic of the α-ketoglutarate co-substrate, inactivates human P4H. The inactivation installs a ketone functionality in P4H, providing a handle for proteomic experiments. Caenorhabditis elegans exposed to the esterified epoxy ketone displays the phenotype of a worm lacking P4H. Thus, this affinity label can be used to mediate collagen stability in an animal, as is desirable in the treatment of a variety of fibrotic diseases. PMID:23702396

  10. 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.

  11. DNA Shape Dominates Sequence Affinity in Nucleosome Formation

    NASA Astrophysics Data System (ADS)

    Freeman, Gordon S.; Lequieu, Joshua P.; Hinckley, Daniel M.; Whitmer, Jonathan K.; de Pablo, Juan J.

    2014-10-01

    Nucleosomes provide the basic unit of compaction in eukaryotic genomes, and the mechanisms that dictate their position at specific locations along a DNA sequence are of central importance to genetics. In this Letter, we employ molecular models of DNA and proteins to elucidate various aspects of nucleosome positioning. In particular, we show how DNA's histone affinity is encoded in its sequence-dependent shape, including subtle deviations from the ideal straight B-DNA form and local variations of minor groove width. By relying on high-precision simulations of the free energy of nucleosome complexes, we also demonstrate that, depending on DNA's intrinsic curvature, histone binding can be dominated by bending interactions or electrostatic interactions. More generally, the results presented here explain how sequence, manifested as the shape of the DNA molecule, dominates molecular recognition in the problem of nucleosome positioning.

  12. Calculation of the Ionization Potentials and Electron Affinities for Atoms

    NASA Astrophysics Data System (ADS)

    Chen, Jiqiang; Krieger, J. B.; Iafrate, G. J.; Savin, A.

    1998-03-01

    The method employing the self-interaction-corrected correlation energy functional obtained from the homogeneous electron gas with a gap is extended to atoms and ions with non-zero spin polarization. As in the case for atoms and ions with ζ=3D0, the error in the calculated Ec is significantly smaller than in the LSD approximation with zero gap for atoms and ions with Z<=18. Comparison of the resulting ionization potentials and electron affinities with experimental values will also be presented. Finally, we will discuss the possibility of obtaining saturation for Ec for the He, Li, N, O, F and Ne isoelectronic series, but a divergent Ec for the Be, B and C isoelectronic series, in the large Z limit.

  13. Scoring Large Scale Affinity Purification Mass Spectrometry Datasets with MIST

    PubMed Central

    Verschueren, Erik; Von Dollen, John; Cimermancic, Peter; Gulbahce, Natali; Sali, Andrej; Krogan, Nevan

    2015-01-01

    High-throughput Affinity Purification Mass Spectrometry (AP-MS) experiments can identify a large number of protein interactions but only a fraction of these interactions are biologically relevant. Here, we describe a comprehensive computational strategy to process raw AP-MS data, perform quality controls and prioritize biologically relevant bait-prey pairs in a set of replicated AP-MS experiments with Mass spectrometry interaction STatistics (MiST). The MiST score is a linear combination of prey quantity (abundance), abundance invariability across repeated experiments (reproducibility), and prey uniqueness relative to other baits (specificity); We describe how to run the full MiST analysis pipeline in an R environment and discuss a number of configurable options that allow the lay user to convert any large-scale AP-MS data into an interpretable, biologically relevant protein-protein interaction network. PMID:25754993

  14. Genetic affinities of Ukrainians from the maternal perspective.

    PubMed

    Pshenichnov, Andrey; Balanovsky, Oleg; Utevska, Olga; Metspalu, Ene; Zaporozhchenko, Valery; Agdzhoyan, Anastasia; Churnosov, Mikhail; Atramentova, Lyubov; Balanovska, Elena

    2013-12-01

    The area of what is now the Ukraine has been the arena of large-scale demographic processes that may have left their traces in the contemporary gene pool of Ukrainians. In this study, we present new mitochondrial DNA data for 607 Ukrainians (hypervariable segment I sequences and coding region polymorphisms). To study the maternal affinities of Ukrainians at the level of separate mitochondrial haplotypes, we apply an original technique, the haplotype co-occurrence analysis. About 20% of the Ukrainian maternal gene pool is represented by lineages highly specific to Ukrainians, but is scarcely found in other populations. About 9% of Ukrainian mtDNA lineages are typical for peoples of the Volga region. We also identified minor gene pool strata (1.6-3.3%), each of which is common in Lithuanians, Estonians, Saami, Nenets, Cornish, and the populations of the North Caucasus.

  15. Political ideology: its structure, functions, and elective affinities.

    PubMed

    Jost, John T; Federico, Christopher M; Napier, Jaime L

    2009-01-01

    Ideology has re-emerged as an important topic of inquiry among social, personality, and political psychologists. In this review, we examine recent theory and research concerning the structure, contents, and functions of ideological belief systems. We begin by defining the construct and placing it in historical and philosophical context. We then examine different perspectives on how many (and what types of) dimensions individuals use to organize their political opinions. We investigate (a) how and to what extent individuals acquire the discursive contents associated with various ideologies, and (b) the social-psychological functions that these ideologies serve for those who adopt them. Our review highlights "elective affinities" between situational and dispositional needs of individuals and groups and the structure and contents of specific ideologies. Finally, we consider the consequences of ideology, especially with respect to attitudes, evaluations, and processes of system justification. PMID:19035826

  16. The metric-affine formalism of f(R) gravity

    NASA Astrophysics Data System (ADS)

    Sotiriou, Thomas P.; Liberati, Stefano

    2007-05-01

    Recently a class of alternative theories of gravity which goes under the name f(R) gravity, has received considerable attention, mainly due to its interesting applications in cosmology. However, the phenomenology of such theories is not only relevant to cosmological scales, especially when it is treated within the framework of the so called Palatini variation, an independent variation with respect to the metric and the connection, which is not considered a priori to be the Levi Civita connection of the metric. If this connection has its standard geometrical meaning the resulting theory will be a metric-affine theory of gravity, as will be discussed in this talk. The general formalism will be presented and several aspects of the theory will be covered, mainly focusing on the enriched phenomenology that such theories exhibit with respect to General Relativity, relevant not only to large scales (cosmology) but also to small scales (e.g. torsion).

  17. Deformation of supersymmetric and conformal quantum mechanics through affine transformations

    NASA Technical Reports Server (NTRS)

    Spiridonov, Vyacheslav

    1993-01-01

    Affine transformations (dilatations and translations) are used to define a deformation of one-dimensional N = 2 supersymmetric quantum mechanics. Resulting physical systems do not have conserved charges and degeneracies in the spectra. Instead, superpartner Hamiltonians are q-isospectral, i.e. the spectrum of one can be obtained from another (with possible exception of the lowest level) by q(sup 2)-factor scaling. This construction allows easily to rederive a special self-similar potential found by Shabat and to show that for the latter a q-deformed harmonic oscillator algebra of Biedenharn and Macfarlane serves as the spectrum generating algebra. A general class of potentials related to the quantum conformal algebra su(sub q)(1,1) is described. Further possibilities for q-deformation of known solvable potentials are outlined.

  18. Search for Amyloid-Binding Proteins by Affinity Chromatography

    PubMed Central

    Calero, Miguel; Rostagno, Agueda; Ghiso, Jorge

    2013-01-01

    ‘Amyloid binging proteins’ is a generic term used to designate proteins that interact with different forms of amyloidogenic peptides or proteins and that, as a result, may modulate their physiological and pathological functions by altering solubility, transport, clearance, degradation, and fibril formation. We describe a simple affinity chromatography protocol to isolate and characterize amyloid-binding proteins based on the use of sequential elution steps that may provide further information on the type of binding interaction. As an example, we depict the application of this protocol to the study of Alzheimer’s amyloid β (Aβ) peptide-binding proteins derived from human plasma. Biochemical analysis of the proteins eluted under different conditions identified serum amyloid P component (SAP) and apolipoprotein J (clusterin) as the main plasma Aβ-binding proteins while various apolipoproteins (apoA-IV, apoE, and apoA-I), as well as albumin (HSA) and fibulin were identified as minor contributors. PMID:22528093

  19. Political ideology: its structure, functions, and elective affinities.

    PubMed

    Jost, John T; Federico, Christopher M; Napier, Jaime L

    2009-01-01

    Ideology has re-emerged as an important topic of inquiry among social, personality, and political psychologists. In this review, we examine recent theory and research concerning the structure, contents, and functions of ideological belief systems. We begin by defining the construct and placing it in historical and philosophical context. We then examine different perspectives on how many (and what types of) dimensions individuals use to organize their political opinions. We investigate (a) how and to what extent individuals acquire the discursive contents associated with various ideologies, and (b) the social-psychological functions that these ideologies serve for those who adopt them. Our review highlights "elective affinities" between situational and dispositional needs of individuals and groups and the structure and contents of specific ideologies. Finally, we consider the consequences of ideology, especially with respect to attitudes, evaluations, and processes of system justification.

  20. Universal vertex-IRF transformation for quantum affine algebras

    SciTech Connect

    Buffenoir, E.; Roche, Ph.; Terras, V.

    2012-10-15

    We construct a universal solution of the generalized coboundary equation in the case of quantum affine algebras, which is an extension of our previous work to U{sub q}(A{sub r}{sup (1)}). This universal solution has a simple Gauss decomposition which is constructed using Sevostyanov's characters of twisted quantum Borel algebras. We show that in the evaluation representations it gives a vertex-face transformation between a vertex type solution and a face type solution of the quantum dynamical Yang-Baxter equation. In particular, in the evaluation representation of U{sub q}(A{sub 1}{sup (1)}), it gives Baxter's well-known transformation between the 8-vertex model and the interaction-round-faces (IRF) height model.

  1. The anatomy, affinity, and phylogenetic significance of Markuelia.

    PubMed

    Dong, Xi-Ping; Donoghue, Philip C J; Cunningham, John A; Liu, Jian-Bo; Cheng, Hong

    2005-01-01

    The fossil record provides a paucity of data on the development of extinct organisms, particularly for their embryology. The recovery of fossilized embryos heralds new insight into the evolution of development but advances are limited by an almost complete absence of phylogenetic constraint. Markuelia is an exception to this, known from cleavage and pre-hatchling stages as a vermiform and profusely annulated direct-developing bilaterian with terminal circumoral and posterior radial arrays of spines. Phylogenetic analyses have hitherto suggested assignment to stem-Scalidophora (phyla Kinorhyncha, Loricifera, Priapulida). We test this assumption with additional data and through the inclusion of additional taxa. The available evidence supports stem-Scalidophora affinity, leading to the conclusion that scalidophorans, cyclonerualians, and ecdysozoans are primitive direct developers, and the likelihood that scalidophorans are primitively metameric. PMID:16174039

  2. The high-affinity maltose switch MBP317-347 has low affinity for glucose: implications for targeting tumors with metabolically directed enzyme prodrug therapy.

    PubMed

    Valdes, Gilmer; Schulte, Reinhard W; Ostermeier, Marc; Iwamoto, Keisuke S

    2014-03-01

    Development of agents with high affinity and specificity for tumor-specific markers is an important goal of molecular-targeted therapy. Here, we propose a shift in paradigm using a strategy that relies on low affinity for fundamental metabolites found in different concentrations in cancerous and non-cancerous tissues: glucose and lactate. A molecular switch, MBP317-347, originally designed to be a high-affinity switch for maltose and maltose-like polysaccharides, was demonstrated to be a low-affinity switch for glucose, that is, able to be activated by high concentrations (tens of millimolar) of glucose. We propose that such a low-affinity glucose switch could be used as a proof of concept for a new prodrug therapy strategy denominated metabolically directed enzyme prodrug therapy (MDEPT) where glucose or, preferably, lactate serves as the activator. Accordingly, considering the typical differential concentrations of lactate found in tumors and in healthy tissues, a low-affinity lactate-binding switch analogous to the low-affinity glucose-binding switch MBP317-347 would be an order of magnitude more active in tumors than in normal tissues and therefore can work as a differential activator of anticancer drugs in tumors.

  3. Proton and sodium cation affinities of harpagide: a computational study.

    PubMed

    Colas, Cyril; Bouchonnet, Stéphane; Rogalewicz-Gilard, Françoise; Popot, Marie-Agnès; Ohanessian, Gilles

    2006-06-15

    The aim of this work was to estimate the proton and sodium cation affinities of harpagide (Har), an iridoid glycoside responsible for the antiinflammatory properties of the medicinal plant Harpagophytum. Monte Carlo conformational searches were performed at the semiempirical AM1 level to determine the most stable conformers for harpagide and its protonated and Na+-cationized forms. The 10 oxygen atoms of the molecule were considered as possible protonation and cationization sites. Geometry optimizations were then refined at the DFT B3LYP/6-31G level from the geometries of the most stable conformers found. Final energetics were obtained at the B3LYP/6-311+G(2d,2p)//B3LYP/6-31G level. The proton and sodium ion affinities of harpagide have been estimated at 223.5 and 66.0 kcal/mol, respectively. Since harpagide mainly provides HarNa+ ions in electrospray experiments, the DeltarG298 associated with the reaction of proton/sodium exchange between Har and methanol, MeOHNa+ + HarH+ --> MeOH2+ + HarNa+ (1), has been calculated; it has been estimated to be 1.9 kcal/mol. Complexing a methanol molecule to each reagent and product of reaction 1 makes the reaction become exothermic by 1.7 kcal/mol. These values are in the limit of the accuracy of the method and do not allow us to conclude definitely whether the reaction is endo- or exothermic, but, according to these very small values, the cation exchange reaction is expected to proceed easily in the final stages of the ion desolvation process. PMID:16759142

  4. Surface Modification of the Conducting Polymer, Polypyrrole, via Affinity Peptide**

    PubMed Central

    Nickels, Jonathan D.; Schmidt, Christine E.

    2012-01-01

    A novel strategy for affinity-based surface modification of the conducting polymer, polypyrrole, (PPy), has been developed. A 12-amino acid peptide (THRTSTLDYFVI, hereafter denoted T59) was previously identified via the phage display technique. This peptide non-covalently binds to the chlorine-doped conducting polymer polypyrrole (PPyCl). Studies have previously shown that conductive polymers have promising application in neural electrodes, sensors, and for improving regeneration and healing of peripheral nerves and other tissues. Thus, the strong and specific attachment of bio-active molecules to the surface of PPy using the T59 affinity peptide is an exciting new approach to enhance the bioactivity of electrically active materials for various biomedical applications. We demonstrate this by using T59 as a tether to modify PPyCl with the laminin fragment IKVAV to enhance cell interactions, as well as with the so-called stealth molecule poly(ethylene glycol; PEG) to decrease cell interactions. Using these two modification strategies, we were able to control cell attachment and neurite extension on the PPy surface, which is critical for different applications (i.e., the goal for tissue regeneration is to enhance cell interactions, whereas the goal for electrode and sensor applications is to reduce glial cell interactions and thus decrease scarring). Significantly, the conductivity of the PPyCl surface was unaffected by this surface modification technique, which is not the case with other methods that have been explored to surface modify conducting polymers. Finally, using subcutaneous implants, we confirmed that the PPyCl treated with the T59 peptide did not react in vivo differently than untreated PPyCl. PMID:23129217

  5. Myo1e binds anionic phospholipids with high affinity.

    PubMed

    Feeser, Elizabeth A; Ignacio, Cherry Mae G; Krendel, Mira; Ostap, E Michael

    2010-11-01

    Myo1e is a single-headed motor protein that has been shown to play roles in clathrin-mediated endocytosis in HeLa cells and podocyte function in the kidney. The myo1e C-terminal tail domain includes a basic region that is required for localization to clathrin-coated vesicles and contains a putative pleckstrin-homology (PH) domain that has been shown to play a role in phospholipid binding in other myosin-I proteins. We used sedimentation assays, stopped-flow fluorescence, and fluorescence microscopy to determine the membrane binding affinities, kinetics, and in vivo localization of fluorescently labeled recombinant myo1e-tail constructs. We found that the myo1e tail binds tightly to large unilamellar vesicles (LUVs) containing physiological concentrations of the anionic phospholipids phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) or phosphatidylserine. The rate of myo1e attachment to LUVs nears the diffusion limit while the calculated rate of detachment from LUVs is slow (k(diss) ≤ 0.4 s(-1)). Mutation of conserved residues in the myo1e PH domain has little effect on lipid binding in vitro or membrane localization in vivo. Soluble inositol phosphate headgroups, such as inositol 1,4,5-trisphosphate, can compete with PtdIns(4,5)P(2) for binding, but the apparent affinity for the soluble inositol phosphate is substantially lower than that for PtdIns(4,5)P(2). These results suggest that myo1e binds lipids through nonspecific electrostatic interactions rather than a stereospecific protein-phosphoinositide interaction. PMID:20860408

  6. Changing water affinity from hydrophobic to hydrophilic in hydrophobic channels.

    PubMed

    Ohba, Tomonori; Yamamoto, Shotaro; Kodaira, Tetsuya; Hata, Kenji

    2015-01-27

    The behavior of water at hydrophobic interfaces can play a significant role in determining chemical reaction outcomes and physical properties. Carbon nanotubes and aluminophosphate materials have one-dimensional hydrophobic channels, which are entirely surrounded by hydrophobic interfaces. Unique water behavior was observed in such hydrophobic channels. In this article, changes in the water affinity in one-dimensional hydrophobic channels were assessed using water vapor adsorption isotherms at 303 K and grand canonical Monte Carlo simulations. Hydrophobic behavior of water adsorbed in channels wider than 3 nm was observed for both adsorption and desorption processes, owing to the hydrophobic environment. However, water showed hydrophilic properties in both adsorption and desorption processes in channels narrower than 1 nm. In intermediate-sized channels, the hydrophobic properties of water during the adsorption process were seen to transition to hydrophilic behavior during the desorption process. Hydrophilic properties in the narrow channels for both adsorption and desorption processes are a result of the relatively strong water-channel interactions (10-15 kJ mol(-1)). In the 2-3 nm channels, the water-channel interaction energy of 4-5 kJ mol(-1) was comparable to the thermal translational energy. The cohesive water interaction was approximately 35 kJ mol(-1), which was larger than the others. Thus, the water affinity change in the 2-3 nm channels for the adsorption and desorption processes was attributed to weak water-channel interactions and strong cohesive interactions. These results are inherently important to control the properties of water in hydrophobic environments.

  7. Brain structure resolves the segmental affinity of anomalocaridid appendages.

    PubMed

    Cong, Peiyun; Ma, Xiaoya; Hou, Xianguang; Edgecombe, Gregory D; Strausfeld, Nicholas J

    2014-09-25

    Despite being among the most celebrated taxa from Cambrian biotas, anomalocaridids (order Radiodonta) have provoked intense debate about their affinities within the moulting-animal clade that includes Arthropoda. Current alternatives identify anomalocaridids as either stem-group euarthropods, crown-group euarthropods near the ancestry of chelicerates, or a segmented ecdysozoan lineage with convergent similarity to arthropods in appendage construction. Determining unambiguous affinities has been impeded by uncertainties about the segmental affiliation of anomalocaridid frontal appendages. These structures are variably homologized with jointed appendages of the second (deutocerebral) head segment, including antennae and 'great appendages' of Cambrian arthropods, or with the paired antenniform frontal appendages of living Onychophora and some Cambrian lobopodians. Here we describe Lyrarapax unguispinus, a new anomalocaridid from the early Cambrian Chengjiang biota, southwest China, nearly complete specimens of which preserve traces of muscles, digestive tract and brain. The traces of brain provide the first direct evidence for the segmental composition of the anomalocaridid head and its appendicular organization. Carbon-rich areas in the head resolve paired pre-protocerebral ganglia at the origin of paired frontal appendages. The ganglia connect to areas indicative of a bilateral pre-oral brain that receives projections from the eyestalk neuropils and compound retina. The dorsal, segmented brain of L. unguispinus reinforces an alliance between anomalocaridids and arthropods rather than cycloneuralians. Correspondences in brain organization between anomalocaridids and Onychophora resolve pre-protocerebral ganglia, associated with pre-ocular frontal appendages, as characters of the last common ancestor of euarthropods and onychophorans. A position of Radiodonta on the euarthropod stem-lineage implies the transformation of frontal appendages to another structure in crown

  8. Rejuvenation of metallic glasses by non-affine thermal strain.

    PubMed

    Ketov, S V; Sun, Y H; Nachum, S; Lu, Z; Checchi, A; Beraldin, A R; Bai, H Y; Wang, W H; Louzguine-Luzgin, D V; Carpenter, M A; Greer, A L

    2015-08-13

    When a spatially uniform temperature change is imposed on a solid with more than one phase, or on a polycrystal of a single, non-cubic phase (showing anisotropic expansion-contraction), the resulting thermal strain is inhomogeneous (non-affine). Thermal cycling induces internal stresses, leading to structural and property changes that are usually deleterious. Glasses are the solids that form on cooling a liquid if crystallization is avoided--they might be considered the ultimate, uniform solids, without the microstructural features and defects associated with polycrystals. Here we explore the effects of cryogenic thermal cycling on glasses, specifically metallic glasses. We show that, contrary to the null effect expected from uniformity, thermal cycling induces rejuvenation, reaching less relaxed states of higher energy. We interpret these findings in the context that the dynamics in liquids become heterogeneous on cooling towards the glass transition, and that there may be consequent heterogeneities in the resulting glasses. For example, the vibrational dynamics of glassy silica at long wavelengths are those of an elastic continuum, but at wavelengths less than approximately three nanometres the vibrational dynamics are similar to those of a polycrystal with anisotropic grains. Thermal cycling of metallic glasses is easily applied, and gives improvements in compressive plasticity. The fact that such effects can be achieved is attributed to intrinsic non-uniformity of the glass structure, giving a non-uniform coefficient of thermal expansion. While metallic glasses may be particularly suitable for thermal cycling, the non-affine nature of strains in glasses in general deserves further study, whether they are induced by applied stresses or by temperature change. PMID:26268190

  9. A High-Affinity Adenosine Kinase from Anopheles Gambiae

    SciTech Connect

    M Cassera; M Ho; E Merino; E Burgos; A Rinaldo-Matthis; S Almo; V Schramm

    2011-12-31

    Genome analysis revealed a mosquito orthologue of adenosine kinase in Anopheles gambiae (AgAK; the most important vector for the transmission of Plasmodium falciparum in Africa). P. falciparum are purine auxotrophs and do not express an adenosine kinase but rely on their hosts for purines. AgAK was kinetically characterized and found to have the highest affinity for adenosine (K{sub m} = 8.1 nM) of any known adenosine kinase. AgAK is specific for adenosine at the nucleoside site, but several nucleotide triphosphate phosphoryl donors are tolerated. The AgAK crystal structure with a bound bisubstrate analogue Ap{sub 4}A (2.0 {angstrom} resolution) reveals interactions for adenosine and ATP and the geometry for phosphoryl transfer. The polyphosphate charge is partly neutralized by a bound Mg{sup 2+} ion and an ion pair to a catalytic site Arg. The AgAK structure consists of a large catalytic core in a three-layer {alpha}/{beta}/{alpha} sandwich, and a small cap domain in contact with adenosine. The specificity and tight binding for adenosine arise from hydrogen bond interactions of Asn14, Leu16, Leu40, Leu133, Leu168, Phe168, and Thr171 and the backbone of Ile39 and Phe168 with the adenine ring as well as through hydrogen bond interactions between Asp18, Gly64, and Asn68 and the ribosyl 2'- and 3'-hydroxyl groups. The structure is more similar to that of human adenosine kinase (48% identical) than to that of AK from Toxoplasma gondii (31% identical). With this extraordinary affinity for AgAK, adenosine is efficiently captured and converted to AMP at near the diffusion limit, suggesting an important role for this enzyme in the maintenance of the adenine nucleotide pool. mRNA analysis verifies that AgAK transcripts are produced in the adult insects.

  10. Brain structure resolves the segmental affinity of anomalocaridid appendages.

    PubMed

    Cong, Peiyun; Ma, Xiaoya; Hou, Xianguang; Edgecombe, Gregory D; Strausfeld, Nicholas J

    2014-09-25

    Despite being among the most celebrated taxa from Cambrian biotas, anomalocaridids (order Radiodonta) have provoked intense debate about their affinities within the moulting-animal clade that includes Arthropoda. Current alternatives identify anomalocaridids as either stem-group euarthropods, crown-group euarthropods near the ancestry of chelicerates, or a segmented ecdysozoan lineage with convergent similarity to arthropods in appendage construction. Determining unambiguous affinities has been impeded by uncertainties about the segmental affiliation of anomalocaridid frontal appendages. These structures are variably homologized with jointed appendages of the second (deutocerebral) head segment, including antennae and 'great appendages' of Cambrian arthropods, or with the paired antenniform frontal appendages of living Onychophora and some Cambrian lobopodians. Here we describe Lyrarapax unguispinus, a new anomalocaridid from the early Cambrian Chengjiang biota, southwest China, nearly complete specimens of which preserve traces of muscles, digestive tract and brain. The traces of brain provide the first direct evidence for the segmental composition of the anomalocaridid head and its appendicular organization. Carbon-rich areas in the head resolve paired pre-protocerebral ganglia at the origin of paired frontal appendages. The ganglia connect to areas indicative of a bilateral pre-oral brain that receives projections from the eyestalk neuropils and compound retina. The dorsal, segmented brain of L. unguispinus reinforces an alliance between anomalocaridids and arthropods rather than cycloneuralians. Correspondences in brain organization between anomalocaridids and Onychophora resolve pre-protocerebral ganglia, associated with pre-ocular frontal appendages, as characters of the last common ancestor of euarthropods and onychophorans. A position of Radiodonta on the euarthropod stem-lineage implies the transformation of frontal appendages to another structure in crown

  11. Rejuvenation of metallic glasses by non-affine thermal strain.

    PubMed

    Ketov, S V; Sun, Y H; Nachum, S; Lu, Z; Checchi, A; Beraldin, A R; Bai, H Y; Wang, W H; Louzguine-Luzgin, D V; Carpenter, M A; Greer, A L

    2015-08-13

    When a spatially uniform temperature change is imposed on a solid with more than one phase, or on a polycrystal of a single, non-cubic phase (showing anisotropic expansion-contraction), the resulting thermal strain is inhomogeneous (non-affine). Thermal cycling induces internal stresses, leading to structural and property changes that are usually deleterious. Glasses are the solids that form on cooling a liquid if crystallization is avoided--they might be considered the ultimate, uniform solids, without the microstructural features and defects associated with polycrystals. Here we explore the effects of cryogenic thermal cycling on glasses, specifically metallic glasses. We show that, contrary to the null effect expected from uniformity, thermal cycling induces rejuvenation, reaching less relaxed states of higher energy. We interpret these findings in the context that the dynamics in liquids become heterogeneous on cooling towards the glass transition, and that there may be consequent heterogeneities in the resulting glasses. For example, the vibrational dynamics of glassy silica at long wavelengths are those of an elastic continuum, but at wavelengths less than approximately three nanometres the vibrational dynamics are similar to those of a polycrystal with anisotropic grains. Thermal cycling of metallic glasses is easily applied, and gives improvements in compressive plasticity. The fact that such effects can be achieved is attributed to intrinsic non-uniformity of the glass structure, giving a non-uniform coefficient of thermal expansion. While metallic glasses may be particularly suitable for thermal cycling, the non-affine nature of strains in glasses in general deserves further study, whether they are induced by applied stresses or by temperature change.

  12. Electron affinity of cubic boron nitride terminated with vanadium oxide

    SciTech Connect

    Yang, Yu; Sun, Tianyin; Shammas, Joseph; Hao, Mei; Nemanich, Robert J.; Kaur, Manpuneet

    2015-10-28

    A thermally stable negative electron affinity (NEA) for a cubic boron nitride (c-BN) surface with vanadium-oxide-termination is achieved, and its electronic structure was analyzed with in-situ photoelectron spectroscopy. The c-BN films were prepared by electron cyclotron resonance plasma-enhanced chemical vapor deposition employing BF{sub 3} and N{sub 2} as precursors. Vanadium layers of ∼0.1 and 0.5 nm thickness were deposited on the c-BN surface in an electron beam deposition system. Oxidation of the metal layer was achieved by an oxygen plasma treatment. After 650 °C thermal annealing, the vanadium oxide on the c-BN surface was determined to be VO{sub 2}, and the surfaces were found to be thermally stable, exhibiting an NEA. In comparison, the oxygen-terminated c-BN surface, where B{sub 2}O{sub 3} was detected, showed a positive electron affinity of ∼1.2 eV. The B{sub 2}O{sub 3} evidently acts as a negatively charged layer introducing a surface dipole directed into the c-BN. Through the interaction of VO{sub 2} with the B{sub 2}O{sub 3} layer, a B-O-V layer structure would contribute a dipole between the O and V layers with the positive side facing vacuum. The lower enthalpy of formation for B{sub 2}O{sub 3} is favorable for the formation of the B-O-V layer structure, which provides a thermally stable surface dipole and an NEA surface.

  13. Development and Characterization of High Affinity Leptins and Leptin Antagonists*

    PubMed Central

    Shpilman, Michal; Niv-Spector, Leonora; Katz, Meirav; Varol, Chen; Solomon, Gili; Ayalon-Soffer, Michal; Boder, Eric; Halpern, Zamir; Elinav, Eran; Gertler, Arieh

    2011-01-01

    Leptin is a pleiotropic hormone acting both centrally and peripherally. It participates in a variety of biological processes, including energy metabolism, reproduction, and modulation of the immune response. So far, structural elements affecting leptin binding to its receptor remain unknown. We employed random mutagenesis of leptin, followed by selection of high affinity mutants by yeast surface display and discovered that replacing residue Asp-23 with a non-negatively charged amino acid leads to dramatically enhanced affinity of leptin for its soluble receptor. Rational mutagenesis of Asp-23 revealed the D23L substitution to be most effective. Coupling the Asp-23 mutation with alanine mutagenesis of three amino acids (L39A/D40A/F41A) previously reported to convert leptin into antagonist resulted in potent antagonistic activity. These novel superactive mouse and human leptin antagonists (D23L/L39A/D40A/F41A), termed SMLA and SHLA, respectively, exhibited over 60-fold increased binding to leptin receptor and 14-fold higher antagonistic activity in vitro relative to the L39A/D40A/F41A mutants. To prolong and enhance in vivo activity, SMLA and SHLA were monopegylated mainly at the N terminus. Administration of the pegylated SMLA to mice resulted in a remarkably rapid, significant, and reversible 27-fold more potent increase in body weight (as compared with pegylated mouse leptin antagonist), because of increased food consumption. Thus, recognition and mutagenesis of Asp-23 enabled construction of novel compounds that induce potent and reversible central and peripheral leptin deficiency. In addition to enhancing our understanding of leptin interactions with its receptor, these antagonists enable in vivo study of the role of leptin in metabolic and immune processes and hold potential for future therapeutic use in disease pathologies involving leptin. PMID:21119198

  14. Development and characterization of high affinity leptins and leptin antagonists.

    PubMed

    Shpilman, Michal; Niv-Spector, Leonora; Katz, Meirav; Varol, Chen; Solomon, Gili; Ayalon-Soffer, Michal; Boder, Eric; Halpern, Zamir; Elinav, Eran; Gertler, Arieh

    2011-02-11

    Leptin is a pleiotropic hormone acting both centrally and peripherally. It participates in a variety of biological processes, including energy metabolism, reproduction, and modulation of the immune response. So far, structural elements affecting leptin binding to its receptor remain unknown. We employed random mutagenesis of leptin, followed by selection of high affinity mutants by yeast surface display and discovered that replacing residue Asp-23 with a non-negatively charged amino acid leads to dramatically enhanced affinity of leptin for its soluble receptor. Rational mutagenesis of Asp-23 revealed the D23L substitution to be most effective. Coupling the Asp-23 mutation with alanine mutagenesis of three amino acids (L39A/D40A/F41A) previously reported to convert leptin into antagonist resulted in potent antagonistic activity. These novel superactive mouse and human leptin antagonists (D23L/L39A/D40A/F41A), termed SMLA and SHLA, respectively, exhibited over 60-fold increased binding to leptin receptor and 14-fold higher antagonistic activity in vitro relative to the L39A/D40A/F41A mutants. To prolong and enhance in vivo activity, SMLA and SHLA were monopegylated mainly at the N terminus. Administration of the pegylated SMLA to mice resulted in a remarkably rapid, significant, and reversible 27-fold more potent increase in body weight (as compared with pegylated mouse leptin antagonist), because of increased food consumption. Thus, recognition and mutagenesis of Asp-23 enabled construction of novel compounds that induce potent and reversible central and peripheral leptin deficiency. In addition to enhancing our understanding of leptin interactions with its receptor, these antagonists enable in vivo study of the role of leptin in metabolic and immune processes and hold potential for future therapeutic use in disease pathologies involving leptin.

  15. Surface modification of the conducting polymer, polypyrrole, via affinity peptide.

    PubMed

    Nickels, Jonathan D; Schmidt, Christine E

    2013-05-01

    A novel strategy for affinity-based surface modification of the conducting polymer, polypyrrole, (PPy), has been developed. A 12-amino acid peptide (THRTSTLDYFVI, hereafter denoted T59) was previously identified via the phage display technique. This peptide noncovalently binds to the chlorine-doped conducting polymer polypyrrole (PPyCl). Studies have previously shown that conductive polymers have promising application in neural electrodes, sensors, and for improving regeneration and healing of peripheral nerves and other tissues. Thus, the strong and specific attachment of bioactive molecules to the surface of PPy using the T59 affinity peptide is an exciting new approach to enhance the bioactivity of electrically active materials for various biomedical applications. We demonstrate this by using T59 as a tether to modify PPyCl with the laminin fragment IKVAV to enhance cell interactions, as well as with the so-called stealth molecule poly(ethylene glycol; PEG) to decrease cell interactions. Using these two modification strategies, we were able to control cell attachment and neurite extension on the PPy surface, which is critical for different applications (i.e., the goal for tissue regeneration is to enhance cell interactions, whereas the goal for electrode and sensor applications is to reduce glial cell interactions and thus decrease scarring). Significantly, the conductivity of the PPyCl surface was unaffected by this surface modification technique, which is not the case with other methods that have been explored to surface modify conducting polymers. Finally, using subcutaneous implants, we confirmed that the PPyCl treated with the T59 peptide did not react in vivo differently than untreated PPyCl. PMID:23129217

  16. SOD1 exhibits allosteric frustration to facilitate metal binding affinity.

    PubMed

    Das, Atanu; Plotkin, Steven S

    2013-03-01

    Superoxide dismutase-1 (SOD1) is a ubiquitous, Cu and Zn binding, free-radical defense enzyme whose misfolding and aggregation play a potential key role in amyotrophic lateral sclerosis, an invariably fatal neurodegenerative disease. Over 150 mutations in SOD1 have been identified with a familial form of the disease, but it is presently not clear what unifying features, if any, these mutants share to make them pathogenic. Here, we develop several unique computational assays for probing the thermo-mechanical properties of both ALS-associated and rationally designed SOD1 variants. Allosteric interaction-free energies between residues and metals are calculated, and a series of atomic force microscopy experiments are simulated with variable tether positions to quantify mechanical rigidity "fingerprints" for SOD1 variants. Mechanical fingerprinting studies of a series of C-terminally truncated mutants, along with an analysis of equilibrium dynamic fluctuations while varying native constraints, potential energy change upon mutation, frustratometer analysis, and analysis of the coupling between local frustration and metal binding interactions for a glycine scan of 90 residues together, reveal that the apo protein is internally frustrated, that these internal stresses are partially relieved by mutation but at the expense of metal-binding affinity, and that the frustration of a residue is directly related to its role in binding metals. This evidence points to apo SOD1 as a strained intermediate with "self-allostery" for high metal-binding affinity. Thus, the prerequisites for the function of SOD1 as an antioxidant compete with apo state thermo-mechanical stability, increasing the susceptibility of the protein to misfold in the apo state.

  17. Cadherin-dependent mechanotransduction depends on ligand identity but not affinity

    PubMed Central

    Tabdili, Hamid; Langer, Matthew; Shi, Quanming; Poh, Yeh-Chuin; Wang, Ning; Leckband, Deborah

    2012-01-01

    Summary This study investigates the relationship between classical cadherin binding affinities and mechanotransduction through cadherin-mediated adhesions. The mechanical properties of cadherin-dependent intercellular junctions are generally attributed to differences in the binding affinities of classical cadherin subtypes that contribute to cohesive energies between cells. However, cell mechanics and mechanotransduction may also regulate intercellular contacts. We used micropipette measurements to quantify the two-dimensional affinities of cadherins at the cell surface, and two complementary mechanical measurements to assess ligand-dependent mechanotransduction through cadherin adhesions. At the cell surface, the classical cadherins investigated in this study form both homophilic and heterophilic bonds with two-dimensional affinities that differ by less than threefold. In contrast, mechanotransduction through cadherin adhesions is strongly ligand dependent such that homophilic, but not heterophilic ligation mediates mechanotransduction, independent of the cadherin binding affinity. These findings suggest that ligand-selective mechanotransduction may supersede differences in cadherin binding affinities in regulating intercellular contacts. PMID:22718345

  18. Purification of proteins specifically binding human endogenous retrovirus K long terminal repeat by affinity elution chromatography.

    PubMed

    Trubetskoy, D O; Zavalova, L L; Akopov, S B; Nikolaev, L G

    2002-11-01

    A novel affinity elution procedure for purification of DNA-binding proteins was developed and employed to purify to near homogeneity the proteins recognizing a 21 base pair sequence within the long terminal repeat of human endogenous retroviruses K. The approach involves loading the initial protein mixture on a heparin-agarose column and elution of protein(s) of interest with a solution of double-stranded oligonucleotide containing binding sites of the protein(s). The affinity elution has several advantages over conventional DNA-affinity chromatography: (i) it is easier and faster, permitting to isolate proteins in a 1 day-one stage procedure; (ii) yield of a target protein is severalfold higher than that in DNA-affinity chromatography; (iii) it is not necessary to prepare a special affinity support for each factor to be isolated. Theaffinity elution could be a useful alternative to conventional DNA-affinity chromatography.

  19. Describing high-dimensional dynamics with low-dimensional piecewise affine models: applications to renewable energy.

    PubMed

    Hirata, Yoshito; Aihara, Kazuyuki

    2012-06-01

    We introduce a low-dimensional description for a high-dimensional system, which is a piecewise affine model whose state space is divided by permutations. We show that the proposed model tends to predict wind speeds and photovoltaic outputs for the time scales from seconds to 100 s better than by global affine models. In addition, computations using the piecewise affine model are much faster than those of usual nonlinear models such as radial basis function models.

  20. [The effect of blood serum polyreactive immunoglobulins on serum antibody affinity determination].

    PubMed

    Bobrovnik, S A; Demchenko, M A; Komisarenko, S V

    2010-01-01

    The presence of polyreactive immunoglobulins in sera may substantially influence on the accuracy of antibody affinity determination. In order to obtain precise values of antibody affinity one should apply one of two following ways. First, one should block polyreactive immunoglobulins with high concentration of Twin 20 and high concentration of any antigen which does not interact with studying antibody. After this antibody affinity may be determined by traditional methods. Another way is the application of the method suggested by us earlier, which allow determining affinity of two antibodies in a mixture and the relation of their concentrations.

  1. Affinity proteomics to study endogenous protein complexes: Pointers, pitfalls, preferences and perspectives

    PubMed Central

    LaCava, John; Molloy, Kelly R.; Taylor, Martin S.; Domanski, Michal; Chait, Brian T.; Rout, Michael P.

    2015-01-01

    Dissecting and studying cellular systems requires the ability to specifically isolate distinct proteins along with the co-assembled constituents of their associated complexes. Affinity capture techniques leverage high affinity, high specificity reagents to target and capture proteins of interest along with specifically associated proteins from cell extracts. Affinity capture coupled to mass spectrometry (MS)-based proteomic analyses has enabled the isolation and characterization of a wide range of endogenous protein complexes. Here, we outline effective procedures for the affinity capture of protein complexes, highlighting best practices and common pitfalls. PMID:25757543

  2. Affinity functions: recognizing essential parameters in fuzzy connectedness based image segmentation

    NASA Astrophysics Data System (ADS)

    Ciesielski, Krzysztof C.; Udupa, Jayaram K.

    2009-02-01

    Fuzzy connectedness (FC) constitutes an important class of image segmentation schemas. Although affinity functions represent the core aspect (main variability parameter) of FC algorithms, they have not been studied systematically in the literature. In this paper, we present a thorough study to fill this gap. Our analysis is based on the notion of equivalent affinities: if any two equivalent affinities are used in the same FC schema to produce two versions of the algorithm, then these algorithms are equivalent in the sense that they lead to identical segmentations. We give a complete characterization of the affinity equivalence and show that many natural definitions of affinity functions and their parameters used in the literature are redundant in the sense that different definitions and values of such parameters lead to equivalent affinities. We also show that two main affinity types - homogeneity based and object feature based - are equivalent, respectively, to the difference quotient of the intensity function and Rosenfeld's degree of connectivity. In addition, we demonstrate that any segmentation obtained via relative fuzzy connectedness (RFC) algorithm can be viewed as segmentation obtained via absolute fuzzy connectedness (AFC) algorithm with an automatic and adaptive threshold detection. We finish with an analysis of possible ways of combining different component affinities that result in non equivalent affinities.

  3. Synthetic studies of neoclerodane diterpenoids from Salvia splendens and evaluation of Opioid Receptor affinity

    PubMed Central

    Fontana, Gianfranco; Savona, Giuseppe; Rodríguez, Benjamín; Dersch, Christina M.; Rothman, Richard B.; Prisinzano, Thomas E.

    2009-01-01

    Salvinorin A (1), a neoclerodane diterpene from the hallucinogenic mint Salvia divinorum, is the only known non-nitrogenous and specific κ-opioid agonist. Several structural congeners of 1 isolated from Salvia splendens (2 – 8) together with a series of semisynthetic derivatives (9 – 24), some of which possess a pyrazoline structural moiety (9, 19 – 22), have been tested for affinity at human μ, δ, and κ opioid receptors. None of these compounds showed high affinity binding to these receptors. However, 10 showed modest affinity for κ receptors suggesting other naturally neoclerodanes from different Salvia species may possess opioid affinity. PMID:20027203

  4. 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

  5. Orientational dependence of the affinity of guanidinium ions to the water surface.

    PubMed

    Wernersson, Erik; Heyda, Jan; Vazdar, Mario; Lund, Mikael; Mason, Philip E; Jungwirth, Pavel

    2011-11-01

    The behavior of guanidinium chloride at the surface of aqueous solutions is investigated using classical molecular dynamics (MD) simulations. It is found that the population of guanidinium ions oriented parallel to the interface is greater in the surface region than in bulk. The opposite is true for ions in other orientations. Overall, guanidinium chloride is depleted in the surface region, in agreement with the fact that the addition of guanidinium chloride increases the surface tension of water. The orientational dependence of the surface affinity of the guanidinium cation is related to its anisotropic hydration. To bring the ion to the surface in the parallel orientation does not require hydrogen bonds to be broken, in contrast to other orientations. The surface enrichment of parallel-oriented guanidinium indicates that its solvation is more favorable near the surface than in bulk solution for this orientation. The dependence of the bulk and surface properties of guanidinium on the force field parameters is also investigated. Despite significant quantitative differences between the force fields, the surface behavior is qualitatively robust. The implications for the orientations of the guanidinium groups of arginine side chains on protein surfaces are also outlined.

  6. Habitat affinity of resident natural enemies of the invasive Aphis glycines (Hemiptera: Aphididae), on soybean, with comments on biological control.

    PubMed

    Brewer, Michael J; Noma, Takuji

    2010-06-01

    We integrated a natural enemy survey of the broader landscape into a more traditional survey for Aphis glycines Matsumura (Hemiptera: Aphididae), parasitoids and predatory flies on soybean using A. glycines-infested soybean, Glycine max (L.) Merr., placed in cropped and noncropped plant systems to complement visual field observations. Across three sites and 5 yr, 18 parasitoids and predatory flies in total (Hymenoptera: Aphelinidae [two species] and Bracondae [seven species], Diptera: Cecidomyiidae [one species], Syrphidae [seven species], Chamaemyiidae [one species]) were detected, with significant variability in recoveries detected across plant system treatments and strong contrasts in habitat affinity detected among species. Lysiphlebus testaceipes Cresson was the most frequently detected parasitoid, and no differences in abundance were detected in cropped (soybean, wheat [Triticum aestivum L.], corn [Zea mays L.], and alfalfa [Medicago sativa L.]) and noncropped (poplar [Populus euramericana (Dode) Guinier] and early successional vegetation) areas. In contrast, Binodoxys kelloggensis Pike, Starý & Brewer had strong habitat affinity for poplar and early successional vegetation. The low recoveries seasonally and across habitats of Aphelinus asychis Walker, Aphelinus sp., and Aphidius colemoni Viereck make their suitability to A. glycines on soybean highly suspect. The widespread occurrence of many of the flies reflects their broad habitat affinity and host aphid ranges. The consistent low field observations of parasitism and predation suggest that resident parasitoids and predatory flies are unlikely to contribute substantially to A. glycines suppression, at least during the conventional time period early in the pest invasion when classical biological control activities are considered. For selected species that were relatively well represented across plant systems (i.e., L. testaceipes and Aphidoletes aphidimyza Rondani), conservation biological control efforts

  7. Electromagnetic field parameters and instrumentation

    NASA Astrophysics Data System (ADS)

    Sheppard, A. R.; Jones, R. A.; Stell, M. E.; Adey, W. R.; Bawin, S.

    1986-07-01

    We studied the effects of the electric and magnetic components of a Loran-C type waveform on three biological systems. Neurochemical assays of brain neurotransmitter substances indicate field-related changes in the levels of norepinephrine in the hippocampus and in the number and affinities of the opiate receptors in the cortex. Behavioral data showed that rats trained in an operant conditioning task did not reliably detect any electric field strength used. Biochemical data demonstrated that the Loran-C field did not modify basal ornithine decarboxylase activity in primary bone cells.

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

    PubMed

    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.

  9. The phylogeny of varanoid lizards and the affinities of snakes

    PubMed Central

    Lee, M. S. Y.

    1997-01-01

    Evidence that platynotan squamates (living varanoid lizards, snakes and their fossil relatives) are monophyletic is presented. Evolutionary relationships within this group are then ascertained through a cladistic analysis of 144 osteological characters. Mosasauroids (aigialosaurs and mosasaurs), a group of large marine lizards, are identified as the nearest relatives of snakes, thus resolving the long-standing problem of snake affinities. The mosasauroid–snake clade (Pythonomorpha) is corroborated by 40 derived characters, including recumbent replacement teeth, thecodonty, four or fewer premaxillary teeth, supratemporal–prootic contact, free mandibular tips, crista circumfenestralis, straight vertical splenio-angular joint, loss of posterior ramus of the coronoid, reduced basipterygoid processes, reduced interpterygoid vacuity, zygosphene–zygantral articulations, and absence of epiphyses on the axial skeleton and skull. After mosasauroids, the next closest relatives of snakes are varanids (Varanus, Saniwa and Saniwides) and lanthanotids (Lanthanotus and Cherminotus). Derived features uniting varanids and lanthanotids include nine cervical vertebrae and three or fewer pairs of sternal ribs. The varanid–lanthanotid–pythonomorph clade, here termed Thecoglossa, is supported by features such as the anteriorly positioned basal tubera, and the loss of the second epibranchial. Successive outgroups to thecoglossans are Telmasaurus, an unresolved polytomy (Estesia, Gobidermatidae and Helodermatidae), Paravaranus and Proplatynota. The 'necrosaurs' are demonstrated to be an artificial (polyphyletic) assemblage of primitive platynotans that are not particularly closely related to each other. Snakes are presumed to have evolved from small, limbless, burrowing lizards and the inability of previous analyses to resolve the affinities of snakes has been attributed to extensive convergence among the numerous lineages of such lizards. The present study contradicts this claim

  10. 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

  11. Transmembrane peptides influence the affinity of sterols for phospholipid bilayers.

    PubMed

    Nyström, Joel H; Lönnfors, Max; Nyholm, Thomas K M

    2010-07-21

    Cholesterol is distributed unevenly between different cellular membrane compartments, and the cholesterol content increases from the inner bilayers toward the plasma membrane. It has been suggested that this cholesterol gradient is important in the sorting of transmembrane proteins. Cholesterol has also been to shown play an important role in lateral organization of eukaryotic cell membranes. In this study the aim was to determine how transmembrane proteins influence the lateral distribution of cholesterol in phospholipid bilayers. Insight into this can be obtained by studying how cholesterol interacts with bilayer membranes of different composition in the presence of designed peptides that mimic the transmembrane helices of proteins. For this purpose we developed an assay in which the partitioning of the fluorescent cholesterol analog CTL between LUVs and mbetaCD can be measured. Comparison of how cholesterol and CTL partitioning between mbetaCD and phospholipid bilayers with different composition suggests that CTL sensed changes in bilayer composition similarly as cholesterol. Therefore, the results obtained with CTL can be used to understand cholesterol distribution in lipid bilayers. The effect of WALP23 on CTL partitioning between DMPC bilayers and mbetaCD was measured. From the results it was clear that WALP23 increased both the order in the bilayers (as seen from CTL and DPH anisotropy) and the affinity of the sterol for the bilayer in a concentration dependent way. Although WALP23 also increased the order in DLPC and POPC bilayers the effects on CTL partitioning was much smaller with these lipids. This indicates that proteins have the largest effect on sterol interactions with phospholipids that have longer and saturated acyl chains. KALP23 did not significantly affect the acyl chain order in the phospholipid bilayers, and inclusion of KALP23 into DMPC bilayers slightly decreased CTL partitioning into the bilayer. This shows that transmembrane proteins can

  12. 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.

  13. C-X...H contacts in biomolecular systems: how they contribute to protein-ligand binding affinity.

    PubMed

    Lu, Yunxiang; Wang, Yong; Xu, Zhijian; Yan, Xiuhua; Luo, Xiaoming; Jiang, Hualiang; Zhu, Weiliang

    2009-09-17

    The hydrogen bond acceptor capability of halogens has long been underappreciated in the field of biology. In this work, we have surveyed structures of protein complexes with halogenated ligands to characterize geometrical preferences of C-X...H contacts and contributions of such interactions to protein-ligand binding affinity. Notably, F...H interactions in biomolecules exhibit a remarkably different behavior as compared to three other kinds of X...H (X = Cl, Br, I) interactions, which has been rationalized by means of ab initio calculations using simple model systems. The C-X...H contacts in biological systems are characterized as weak hydrogen bonding interactions. Furthermore, the electrophile "head on" and nucleophile "side on" interactions of halogens have been extensively investigated through the examination of interactions in protein structures and a two-layer ONIOM-based QM/MM method. In biomolecular systems, C-X...H contacts are recognized as secondary interaction contributions to C-X...O halogen bonds that play important roles in conferring specificity and affinity for halogenated ligands. The results presented here are within the context of their potential applications in drug design, including relevance to the development of accurate force fields for halogens.

  14. Using three-dimensional quantitative structure-activity relationships to examine estrogen receptor binding affinities of polychlorinated hydroxybiphenyls

    SciTech Connect

    Waller, C.L.; Minor, D.L.; McKinney, J.D.

    1995-07-01

    Certain phenyl-substituted hydrocarbons of environmental concern have the potential to disrupt the endocrine system of animals, apparently in association with their estrogenic properties. Competition with natural estrogens for the estrogen receptor is a possible mechanism by which such effects could occur. We used comparative molecular field analysis (CoMFA), a three-dimensional quantitative structure-activity relationship (QSAR) paradigm, to examine the underlying structural properties of ortho-chlorinated hydroxybiphenyl analogs known to bind to the estrogen receptor. The cross-validated and conventional statistical results indicate a high degree of internal predictability for the molecules included in the training data set. In addition to the phenolic (A) ring system, conformational restriction of the overall structure appears to play an important role in estrogen receptor binding affinity. Hydrophobic character as assessed using hydropathic interaction fields also contributes in a positive way to binding affinity. The CoMFA-derived QSARs may be useful in examining the estrogenic activity of a wider range of phenyl-substituted hydrocarbons of environmental concern. 37 refs., 2 figs., 2 tabs.

  15. Jen1p: A High Affinity Selenite Transporter in Yeast

    PubMed Central

    McDermott, Joseph R.; Rosen, Barry P.

    2010-01-01

    Selenium is a micronutrient in most eukaryotes, including humans, which is well known for having an extremely thin border between beneficial and toxic concentrations. Soluble tetravalent selenite is the predominant environmental form and also the form that is applied in the treatment of human diseases. To acquire this nutrient from low environmental concentrations as well as to avoid toxicity, a well-controlled transport system is required. Here we report that Jen1p, a proton-coupled monocarboxylate transporter in S. cerevisiae, catalyzes high-affinity uptake of selenite. Disruption of JEN1 resulted in selenite resistance, and overexpression resulted in selenite hypersensitivity. Transport assay showed that overexpression of Jen1p enables selenite accumulation in yeast compared with a JEN1 knock out strain, indicating the Jen1p transporter facilitates selenite accumulation inside cells. Selenite uptake by Jen1p had a Km of 0.91 mM, which is comparable to the Km for lactate. Jen1p transported selenite in a proton-dependent manner which resembles the transport mechanism for lactate. In addition, selenite and lactate can inhibit the transport of each other competitively. Therefore, we postulate selenite is a molecular mimic of monocarboxylates which allows selenite to be transported by Jen1p. PMID:20861301

  16. Preparation of phenylboronate affinity rigid monolith with macromolecular porogen.

    PubMed

    Li, Xiang-Jie; Jia, Man; Zhao, Yong-Xin; Liu, Zhao-Sheng; Akber Aisa, Haji

    2016-03-18

    Boronate-affinity monolithic column was first prepared via polystyrene (PS) as porogen in this work. The monolithic polymer was synthetized using 4-vinylphenylboronic acid (4-VPBA) as functional monomer, ethylene glycol dimethacrylate (EDMA) as crosslinker monomer, and a mixture of PS solution in tetrahydrofuran, the linear macromolecular porogen, and toluene as porogen. Isoquercitrin (ISO) and hyperoside (HYP), isomer diol flavonoid glycosides, can be baseline separated on the poly(VPBA-co-EDMA) monolith. The effect of polymerization variables on the selectivity factor, e.g., the ratio of monomer to crosslinker (M/C), the amount of PS and the molecular weight of macromolecular porogen was investigated. The surface properties of the monolithic polymer were characterized by scanning electron microscopy and nitrogen adsorption. The best polymerization condition was the M/C ratio of 7:3, and the PS concentration of 40 mg/ml. The poly(VPBA-co-EDMA) polymer was also applied to extract cis-diol flavonoid glycosides from the crude extraction of cotton flower. After treated by poly(VPBA-co-EDMA) for solid phase extraction, high purity ISO and HYP (>99.96%) can be obtained with recovery of 83.7% and 78.6%, respectively. PMID:26896914

  17. Methods for determining the genetic affinity of microorganisms and viruses

    NASA Technical Reports Server (NTRS)

    Fox, George E. (Inventor); Willson, III, Richard C. (Inventor); Zhang, Zhengdong (Inventor)

    2012-01-01

    Selecting which sub-sequences in a database of nucleic acid such as 16S rRNA are highly characteristic of particular groupings of bacteria, microorganisms, fungi, etc. on a substantially phylogenetic tree. Also applicable to viruses comprising viral genomic RNA or DNA. A catalogue of highly characteristic sequences identified by this method is assembled to establish the genetic identity of an unknown organism. The characteristic sequences are used to design nucleic acid hybridization probes that include the characteristic sequence or its complement, or are derived from one or more characteristic sequences. A plurality of these characteristic sequences is used in hybridization to determine the phylogenetic tree position of the organism(s) in a sample. Those target organisms represented in the original sequence database and sufficient characteristic sequences can identify to the species or subspecies level. Oligonucleotide arrays of many probes are especially preferred. A hybridization signal can comprise fluorescence, chemiluminescence, or isotopic labeling, etc.; or sequences in a sample can be detected by direct means, e.g. mass spectrometry. The method's characteristic sequences can also be used to design specific PCR primers. The method uniquely identifies the phylogenetic affinity of an unknown organism without requiring prior knowledge of what is present in the sample. Even if the organism has not been previously encountered, the method still provides useful information about which phylogenetic tree bifurcation nodes encompass the organism.

  18. Solubilization and partial characterization of a microsomal high affinity GTPase

    SciTech Connect

    Nicchitta, C.; Williamson, J.R.

    1987-05-01

    Isolated rat liver microsomes release sequestered Ca/sup 2 +/ following addition of GTP. In contrast to permeabilized cells, GTP dependent microsomal Ca/sup 2 +/ release requires low concentrations of polyethylene glycol (PEG). They have identified a microsomal, PEG-sensitive high affinity GTPase which shares a number of characteristics with the GTP-dependent Ca/sup 2 +/ release system. To aid in further characterization of this activity they have initiated studies on the solubilization and purification of the microsomal GTPases. When microsomes are solubilized under the following conditions (150 mM NaCl, 5 mg protein/ml, 1% Triton X-114) PEG sensitive GTPase activity selectively partitions into the detergent rich phase of the Triton X-114 extract. As observed in intact microsomal membranes the Triton X-114 soluble GTPase is maximally stimulated by 3% PEG. Half maximal stimulation is observed at 1% PEG. PEG increases the Vmax of this activity; no effects on Km were observed. The Km for GTP of the detergent soluble GTPase is 5 ..mu..M. This GTPase is sensitive to inhibition by sulfhydryl reagents. PEG-sensitive GTPase activity was completely inhibited in the presence of 25 ..mu..M p-hydroxymercuribenzoate (PHMB); half maximal inhibition was observed at 5 ..mu..M. Labeling of the Triton X-114 extract with the photosensitive compound (/sup 32/P) 8-azido GTP indicated the presence of two prominent GTP binding proteins of approximate molecular weights 17 and 54 kD.

  19. Magnetic particles as affinity matrix for purification of antithrombin

    NASA Astrophysics Data System (ADS)

    Mercês, A. A. D.; Maciel, J. C.; Carvalho Júnior, L. B.

    2015-11-01

    Immobilization of biomolecules onto insoluble supports is an important tool for the fabrication of a diverse range of functional materials. It provides advantages: enhanced stability and easy separation. In this work two different magnetic composites were synthesized (MAG-PANI-HS and mDAC-HS) to human antithrombin purification. The magnetic particles (MAG) were obtained by co-precipitation method of iron salts II and III and subsequently coated with polyaniline (MAG-PANI particles). Dacron (polyethylene terephthalate) suffered a hydrazinolysis reaction to obtain a powder (Dacron hydrazide) which was subsequently magnetized (mDAC particles) also by co-precipitation method. Heparan sulfate (HS) was immobilized to MAG-PANI and mDAC retained respectively 35μg and 38.6μg per of support. The magnetic composite containing HS immobilized (MAG-PANI-HS and mDAC-HS) was incubated with human blood plasma (1mL) and then washed with NaCl gradients. Electrophoresis of proteins present in eluates showed bands of antithrombin (58kDa). A reduction in the antithrombin activity was detected in plasma that were incubated in the composites magnetic with HS immobilized, suggesting that the antithrombin was removed of the human blood plasma and then purified. Therefore, the above results suggest that both preparations: MAG-PANI-HS and mDAC-HS are able to affinity purify antithrombin, an important component of blood coagulation.

  20. Affinity and diversity indices for anopheline immature forms.

    PubMed

    Nagm, Lucy; Luitgards-Moura, José Francisco; Neucamp, César de Souza; Monteiro-de-Barros, Fábio Saito; Honório, Nildimar Alves; Tsouris, Pantelis; Rosa-Freitas, Maria Goreti

    2007-01-01

    As for the entire Amazon Region, malaria continues to be a major health public problem in Roraima that presented an Annual Parasitic Index of 85.4 in 2005, the highest in Brazil. Information on anopheline breeding sites is an essential component in malaria control strategies. Aiming to contribute to the limited knowledge on anopheline immature forms in Roraima, collections and breeding site observations were performed in 10 breeding sites around the capital city Boa Vista. Collections were carried out in the rainy and dry season periods between April 2004 and January 2005. Breeding sites comprised natural and artificial water reservoirs. A total of 623 immature forms were collected belonging to Anopheles albitarsis s.l., An.triannulatus s.l., An. nuneztovari/dunhami, An. braziliensis, An. evansae, An. oswaldoi s.l., An. strodei and An. darlingi. An. albitarsis and An. braziliensis were the most frequently found species. Eight larvae of An. darlingi were found in only one breeding site located in the forest. An. triannulatus/An. nuneztovari and An. albitarsis/An. braziliensis were the pairs of species that mostly occurred together. Both pair of species displayed the highest affinity index what might indicate a high compatibility for the same breeding conditions and/or a synergistic co-occurrence. Species diversity index was higher for the dry season.

  1. Phosphatidylserine Reversibly Binds Cu2+ with Extremely High Affinity

    PubMed Central

    Monson, Christopher F.; Cong, Xiao; Robison, Aaron; Pace, Hudson P.; Liu, Chunming; Poyton, Matthew F.; Cremer, Paul S.

    2012-01-01

    Phosphatidylserine (PS) embedded within supported lipid bilayers (SLBs) was found to bind Cu2+ from solution with extraordinarily high affinity. In fact, the equilibrium dissociation constant was in the femtomolar range. The resulting complex formed in a 1:2 Cu2+ to PS ratio and quenches a broad spectrum of lipid-bound fluorophores in a reversible and pH-dependent fashion. At acidic pH values, the fluorophores were almost completely unquenched, while at basic pH values significant quenching (85–90%) was observed. The pH at which the transition occurred was dependent on the PS concentration and ranged from approximately pH 5 to 8. The quenching kinetics was slow at low Cu2+ concentrations and basic values pH (up to several hours), while the unquenching reaction was orders of magnitude more rapid upon lowering the pH. This was consistent with diffusion limited complex formation at basic pH, but rapid dissociation under acidic conditions. The tight binding of Cu2+ to PS may have physiological consequences under certain circumstances. PMID:22548290

  2. Preparation of phenylboronate affinity rigid monolith with macromolecular porogen.

    PubMed

    Li, Xiang-Jie; Jia, Man; Zhao, Yong-Xin; Liu, Zhao-Sheng; Akber Aisa, Haji

    2016-03-18

    Boronate-affinity monolithic column was first prepared via polystyrene (PS) as porogen in this work. The monolithic polymer was synthetized using 4-vinylphenylboronic acid (4-VPBA) as functional monomer, ethylene glycol dimethacrylate (EDMA) as crosslinker monomer, and a mixture of PS solution in tetrahydrofuran, the linear macromolecular porogen, and toluene as porogen. Isoquercitrin (ISO) and hyperoside (HYP), isomer diol flavonoid glycosides, can be baseline separated on the poly(VPBA-co-EDMA) monolith. The effect of polymerization variables on the selectivity factor, e.g., the ratio of monomer to crosslinker (M/C), the amount of PS and the molecular weight of macromolecular porogen was investigated. The surface properties of the monolithic polymer were characterized by scanning electron microscopy and nitrogen adsorption. The best polymerization condition was the M/C ratio of 7:3, and the PS concentration of 40 mg/ml. The poly(VPBA-co-EDMA) polymer was also applied to extract cis-diol flavonoid glycosides from the crude extraction of cotton flower. After treated by poly(VPBA-co-EDMA) for solid phase extraction, high purity ISO and HYP (>99.96%) can be obtained with recovery of 83.7% and 78.6%, respectively.

  3. 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

  4. Limited proteolysis for assaying ligand binding affinities of nuclear receptors.

    PubMed

    Benkoussa, M; Nominé, B; Mouchon, A; Lefebvre, B; Bernardon, J M; Formstecher, P; Lefebvre, P

    1997-01-01

    The binding of natural or synthetic ligands to nuclear receptors is the triggering event leading to gene transcription activation or repression. Ligand binding to the ligand binding domain of these receptors induces conformational changes that are evidenced by an increased resistance of this domain to proteases. In vitro labeled receptors were incubated with various synthetic or natural agonists or antagonists and submitted to trypsin digestion. Proteolysis products were separated by SDS-PAGE and quantified. The amount of trypsin-resistant fragments was proportional to receptor occupancy by the ligand, and allowed the determination of dissociation constants (kDa). Using the wild-type or mutated human retinoic acid receptor alpha as a model, kDa values determined by classical competition binding assays using tritiated ligands are in agreement with those measured by the proteolytic assay. This method was successfully extended to human retinoic X receptor alpha, glucocorticoid receptor, and progesterone receptor, thus providing a basis for a new, faster assay to determine simultaneously the affinity and conformation of receptors when bound to a given ligand.

  5. Sliding Window Generalized Kernel Affine Projection Algorithm Using Projection Mappings

    NASA Astrophysics Data System (ADS)

    Slavakis, Konstantinos; Theodoridis, Sergios

    2008-12-01

    Very recently, a solution to the kernel-based online classification problem has been given by the adaptive projected subgradient method (APSM). The developed algorithm can be considered as a generalization of a kernel affine projection algorithm (APA) and the kernel normalized least mean squares (NLMS). Furthermore, sparsification of the resulting kernel series expansion was achieved by imposing a closed ball (convex set) constraint on the norm of the classifiers. This paper presents another sparsification method for the APSM approach to the online classification task by generating a sequence of linear subspaces in a reproducing kernel Hilbert space (RKHS). To cope with the inherent memory limitations of online systems and to embed tracking capabilities to the design, an upper bound on the dimension of the linear subspaces is imposed. The underlying principle of the design is the notion of projection mappings. Classification is performed by metric projection mappings, sparsification is achieved by orthogonal projections, while the online system's memory requirements and tracking are attained by oblique projections. The resulting sparsification scheme shows strong similarities with the classical sliding window adaptive schemes. The proposed design is validated by the adaptive equalization problem of a nonlinear communication channel, and is compared with classical and recent stochastic gradient descent techniques, as well as with the APSM's solution where sparsification is performed by a closed ball constraint on the norm of the classifiers.

  6. Two measured completely different electron affinities for atomic Eu?

    NASA Astrophysics Data System (ADS)

    Msezane, A. Z.; Felfli, Z.

    2016-05-01

    Recently, the electron affinity (EA) of atomic Eu was measured to be 0.116?eV. This value is in outstanding agreement with the theoretically calculated values using the Regge pole and MCDF-RCI methods. Previously, the EA of Eu was measured to be 1.053 eV. In an attempt to resolve the discrepancy between the two measured values, we have adopted the complex angular momentum (CAM) method and investigated in the electron energy range 0.11 eV

  7. An affinity-directed protein missile system for targeted proteolysis

    PubMed Central

    Fulcher, Luke J.; Macartney, Thomas; Bozatzi, Polyxeni; Hornberger, Annika; Rojas-Fernandez, Alejandro

    2016-01-01

    The von Hippel–Lindau (VHL) protein serves to recruit the hypoxia-inducible factor alpha (HIF1α) protein under normoxia to the CUL2 E3 ubiquitin ligase for its ubiquitylation and degradation through the proteasome. In this report, we modify VHL to engineer an affinity-directed protein missile (AdPROM) system to direct specific endogenous target proteins for proteolysis in mammalian cells. The proteolytic AdPROM construct harbours a cameloid anti-green fluorescence protein (aGFP) nanobody that is fused to VHL for either constitutive or tetracycline-inducible expression. For target proteins, we exploit CRISPR/Cas9 to rapidly generate human kidney HEK293 and U2OS osteosarcoma homozygous knock-in cells harbouring GFP tags at the VPS34 (vacuolar protein sorting 34) and protein associated with SMAD1 (PAWS1, aka FAM83G) loci, respectively. Using these cells, we demonstrate that the expression of the VHL-aGFP AdPROM system results in near-complete degradation of the endogenous GFP-VPS34 and PAWS1-GFP proteins through the proteasome. Additionally, we show that Tet-inducible destruction of GFP-VPS34 results in the degradation of its associated partner, UVRAG, and reduction in levels of cellular phosphatidylinositol 3-phosphate. PMID:27784791

  8. Ethanol increases affinity of protein kinase C for phosphatidylserine

    SciTech Connect

    Chin, J.H.

    1986-03-01

    Protein kinase C is a calcium-dependent enzyme that requires phospholipid for its activation. It is present in relatively high concentration in the brain and may be involved in neuronal function. The present experiments test whether the membrane disorder induced by ethanol affects the activity of kinase C by changing its interaction with membrane lipid. Fractions rich in kinase C were purified from rat brain cytosol by DEAE-cellulose chromatography and Sephadex G-200 gel filtration. Enzyme activity was assayed by measuring the phosphorylation of histone H1. As expected, phosphatidylserine activated the enzyme, and the stimulation was further increased by the addition of calcium and/or diacylglycerol. At low concentration of free calcium (0.5-1..mu..M), ethanol (800 mM0 enhanced kinase C activity if the presence of phospholipid. similar results were observed in the absence of calcium. Double reciprocal plots of the data showed that ethanol increased the affinity of the enzyme for phosphatidylserine without affecting the V/sub max. The stimulation of kinase C activity by ethanol was not observed at high calcium concentrations. These experiments suggest that ethanol may activated protein kinase C at physiological levels of calcium by facilitating its transfer into the hydrophobic membrane environment.

  9. Production and Purification of Streptokinase by Protected Affinity Chromatography

    PubMed Central

    Babashamsi, Mohammad; Razavian, Mohammad Hossein; Nejadmoghaddam, Mohammad Reza

    2009-01-01

    Streptokinase is an extracellular protein, extracted from certain strains of beta hemolytic streptococcus. It is a non-protease plasminogen activator that activates plasminogen to plasmin, the enzyme that degrades fibrin cloth through its specific lysine binding site; it is used therefore as a drug in thrombolytic therapy. The rate of bacterial growth and streptokinase production was studied in condition of excess glucose addition to culture media and its pH maintenance. The streptokinase product of the bacterial culture was preliminary extracted by salt precipitation and then purified by affinity chromatography on plasminogen substituted sepharose-4B in a condition that the plasminogen active site was protected from streptokinase-induced activation. The purity of streptokinase was confirmed by SDS-PAGE and its biological activity determined in a specific streptokinase assay. The results showed that in the fed–batch culture, the rate of streptokinase production increased over two times as compared with the batch culture while at the same time, shortening the streptokinase purification to a single step increased the yield over 95% at the chromatography stage. PMID:23407807

  10. Mining the soluble chloroplast proteome by affinity chromatography

    PubMed Central

    Bayer, Roman G; Stael, Simon; Csaszar, Edina; Teige, Markus

    2011-01-01

    Chloroplasts are fundamental organelles enabling plant photoautotrophy. Besides their outstanding physiological role in fixation of atmospheric CO2, they harbor many important metabolic processes such as biosynthesis of amino acids, vitamins or hormones. Technical advances in MS allowed the recent identification of most chloroplast proteins. However, for a deeper understanding of chloroplast function it is important to obtain a complete list of constituents, which is so far limited by the detection of low-abundant proteins. Therefore, we developed a two-step strategy for the enrichment of low-abundant soluble chloroplast proteins from Pisum sativum and their subsequent identification by MS. First, chloroplast protein extracts were depleted from the most abundant protein ribulose-1,5-bisphosphate carboxylase/oxygenase by SEC or heating. Further purification was carried out by affinity chromatography, using ligands specific for ATP- or metal-binding proteins. By these means, we were able to identify a total of 448 proteins including 43 putative novel chloroplast proteins. Additionally, the chloroplast localization of 13 selected proteins was confirmed using yellow fluorescent protein fusion analyses. The selected proteins included a phosphoglycerate mutase, a cysteine protease, a putative protein kinase and an EF-hand containing substrate carrier protein, which are expected to exhibit important metabolic or regulatory functions. PMID:21365755

  11. A designed repeat protein as an affinity capture reagent.

    PubMed

    Speltz, Elizabeth B; Brown, Rebecca S H; Hajare, Holly S; Schlieker, Christian; Regan, Lynne

    2015-10-01

    Repeat proteins are an attractive target for protein engineering and design. We have focused our attention on the design and engineering of one particular class: tetratricopeptide repeat (TPR) proteins. In previous work, we have shown that the structure and stability of TPR proteins can be manipulated in a rational fashion [Cortajarena (2011) Prot. Sci. 20: , 1042-1047; Main (2003) Structure 11: , 497-508]. Building on those studies, we have designed and characterized a number of different peptide-binding TPR modules and we have also assembled these modules into supramolecular arrays [Cortajarena (2009) ACS Chem. Biol. 5: , 545-552; Cortajarena (2008) ACS Chem. Biol. 3: , 161-166; Jackrel (2009) Prot. Sci. 18: , 762-774; Kajander (2007) Acta Crystallogr. D Biol. Crystallogr. 63: , 800-811]. Here we focus on the development of one such TPR-peptide interaction for a practical application, affinity purification. We illustrate the general utility of our designed protein interaction. Furthermore, this example highlights how basic research on protein-peptide interactions can lead to the development of novel reagents with important practical applications. PMID:26517897

  12. Defining the human gallbladder proteome by transcriptomics and affinity proteomics.

    PubMed

    Kampf, Caroline; Mardinoglu, Adil; Fagerberg, Linn; Hallström, Björn M; Danielsson, Angelika; Nielsen, Jens; Pontén, Fredrik; Uhlen, Mathias

    2014-11-01

    Global protein analysis of human gallbladder tissue is vital for identification of molecular regulators and effectors of its physiological activity. Here, we employed a genome-wide deep RNA sequencing analysis in 28 human tissues to identify the genes overrepresented in the gallbladder and complemented it with antibody-based immunohistochemistry in 48 human tissues. We characterized human gallbladder proteins and identified 140 gallbladder-specific proteins with an elevated expression in the gallbladder as compared to the other analyzed tissues. Five genes were categorized as enriched, with at least fivefold higher levels in gallbladder, 60 genes were categorized as group enriched with elevated transcript levels in gallbladder shared with at least one other tissue and 75 genes were categorized as enhanced with higher expression than the average expression in other tissues. We explored the localization of the genes within the gallbladder through cell-type specific antibody-based protein profiling and the subcellular localization of the genes through immunofluorescent-based profiling. Finally, we revealed the biological processes and metabolic functions carried out by these genes through the use of GO, KEGG Pathway, and HMR2.0 that is compilation of the human metabolic reactions. We demonstrated the results of the combined analysis of the transcriptomics and affinity proteomics.

  13. Proteomic analysis and discovery using affinity proteomics and mass spectrometry.

    PubMed

    Olsson, Niclas; Wingren, Christer; Mattsson, Mikael; James, Peter; O'Connell, David; Nilsson, Fredrik; Cahill, Dolores J; Borrebaeck, Carl A K

    2011-10-01

    Antibody-based microarrays are a rapidly evolving affinity-proteomic methodology that recently has shown great promise in clinical applications. The resolution of these proteomic analyses is, however, directly related to the number of data-points, i.e. antibodies, included on the array. Currently, this is a key bottleneck because of limited availability of numerous highly characterized antibodies. Here, we present a conceptually new method, denoted global proteome survey, opening up the possibility to probe any proteome in a species-independent manner while still using a limited set of antibodies. We use context-independent-motif-specific antibodies directed against short amino acid motifs, where each motif is present in up to a few hundred different proteins. First, the digested proteome is exposed to these antibodies, whereby motif-containing peptides are enriched, which then are detected and identified by mass spectrometry. In this study, we profiled extracts from human colon tissue, yeast cells lysate, and mouse liver tissue to demonstrate proof-of-concept.

  14. Labeling proteins by affinity-guided DMAP chemistry.

    PubMed

    Tamura, Tomonori; Hamachi, Itaru

    2015-01-01

    Catalysts have long played an essential role in organic synthesis and thus hold potential as tools for chemical protein modification. However, there are only a few examples of catalyst-mediated protein labeling under biological conditions because of the difficulty of designing molecular catalysts that work in aqueous environments with high target selectivity and reaction efficiency. To overcome this situation, we have previously developed a new catalyst-based method, termed affinity-guided DMAP (4-dimethylaminopyridine) (AGD) chemistry, for site-specific protein labeling in a target-selective manner using an acyl transfer reaction. More recently, we discovered that the labeling rate and efficiency can be greatly enhanced by using "multivalent" DMAP groups. Here, we describe the principle of the multivalent AGD chemistry and the protocol for chemical labeling of FK506-binding protein 12 (FKBP12) in test tubes. In this method, the FKBP12 labeling is completed within 30 min and occurs site specifically at the vicinity of the ligand-binding pocket of the protein.

  15. Aptamer stationary phase for protein capture in affinity capillary chromatography.

    PubMed

    Connor, Adam C; McGown, Linda B

    2006-04-14

    The thrombin-binding DNA aptamer was used with thrombin as a model system to investigate protein capture using aptamer stationary phases in affinity capillary chromatography. The aptamer was covalently attached to the inner surface of a bare fused-silica glass capillary to serve as the stationary phase. Proteins were loaded onto the capillary via an applied pressure. The capillary was then washed to remove unbound and non-specifically associated proteins. Finally, the bound protein was released and eluted using 20 mM Tris buffer containing 8 M urea, pH 7.3, at 50 degrees C. Eluate was collected after each step (load, wash and elute) and relative amounts of protein each were compared using fluorescence spectroscopy. The identity of the protein in the collections was confirmed using matrix assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry. The experiment was repeated for thrombin on a bare (unmodified) capillary and a capillary coated with a scrambled-sequence, non-G-quartet forming oligonucleotide that does not bind with thrombin. The results show that the aptamer stationary phase captures approximately three times as much thrombin as the control columns. The experiment was also repeated using human serum albumin (HSA) alone and in an equimolar mixture with thrombin. HSA was not retained on the aptamer capillary, nor did it affect the capture of thrombin from the mixture.

  16. Generation of high-performance binding proteins for peptide motifs by affinity clamping

    PubMed Central

    Koide, Shohei; Huang, Jin

    2013-01-01

    We describe concepts and methodologies for generating “Affinity Clamps”, a new class of recombinant binding proteins that achieve high affinity and high specificity toward short peptide motifs of biological importance, which is a major challenge in protein engineering. The Affinity Clamping concept exploits the potential of nonhomologous recombination of protein domains in generating large changes in protein function and the inherent binding affinity and specificity of the so-called modular interaction domains toward short peptide motifs. Affinity Clamping creates a clamshell architecture that clamps onto a target peptide. The design processes involve (i) choosing a starting modular interaction domain appropriate for the target and applying structure-guided modifications, (ii) attaching a second domain, termed “enhancer domain” and (iii) optimizing the peptide-binding site located between the domains by directed evolution. The two connected domains work synergistically to achieve high levels of affinity and specificity that are unattainable with either domain alone. Because of the simple and modular architecture, affinity clamps are particularly well suited as building blocks for designing more complex functionalities. Affinity Clamping represents a major advance in protein design that is broadly applicable to the recognition of peptide motifs. PMID:23422435

  17. Serial studies on the affinity and heterogeneity of human autoantibodies to thyroglobulin.

    PubMed Central

    Devey, M E; Bleasdale-Barr, K M; McLachlan, S M; Bradbury, J; Clark, F; Young, E T

    1989-01-01

    The functional affinity and heterogeneity of autoantibodies to thyroglobulin (Tg) were measured by an IgG subclass-specific solid-phase competition ELISA in patients with autoimmune thyroid disease. High-affinity IgG1 and IgG4 antibodies formed the major anti-Tg response. Both titre and affinity of IgG3 and IgG2 anti-Tg were generally low but in some Hashimoto's disease patients high-affinity IgG2 anti-Tg were found and IgG2 anti-Tg, unlike those of other subclasses, showed very restricted heterogeneity. The affinity of IgG4 anti-Tg was similar in patients with thyroid disease and their clinically euthyroid (normal) relatives. In contrast, a progressive increase in IgG1 anti-Tg affinity was seen in clinically euthyroid individuals compared with their relatives with thyroid disease and high titred Hashimoto's disease patients, suggesting that either rising titres of high affinity IgG1 anti-Tg or affinity maturation of IgG1 anti-Tg may be indicative of impending hypothyroidism. PMID:2776357

  18. 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...

  19. Structural Insights into the Affinity of Cel7A Carbohydrate-binding Module for Lignin*

    PubMed Central

    Strobel, Kathryn L.; Pfeiffer, Katherine A.; Blanch, Harvey W.; Clark, Douglas S.

    2015-01-01

    The high cost of hydrolytic enzymes impedes the commercial production of lignocellulosic biofuels. High enzyme loadings are required in part due to their non-productive adsorption to lignin, a major component of biomass. Despite numerous studies documenting cellulase adsorption to lignin, few attempts have been made to engineer enzymes to reduce lignin binding. In this work, we used alanine-scanning mutagenesis to elucidate the structural basis for the lignin affinity of Trichoderma reesei Cel7A carbohydrate binding module (CBM). T. reesei Cel7A CBM mutants were produced with a Talaromyces emersonii Cel7A catalytic domain and screened for their binding to cellulose and lignin. Mutation of aromatic and polar residues on the planar face of the CBM greatly decreased binding to both cellulose and lignin, supporting the hypothesis that the cellulose-binding face is also responsible for lignin affinity. Cellulose and lignin affinity of the 31 mutants were highly correlated, although several mutants displayed selective reductions in lignin or cellulose affinity. Four mutants with increased cellulose selectivity (Q2A, H4A, V18A, and P30A) did not exhibit improved hydrolysis of cellulose in the presence of lignin. Further reduction in lignin affinity while maintaining a high level of cellulose affinity is thus necessary to generate an enzyme with improved hydrolysis capability. This work provides insights into the structural underpinnings of lignin affinity, identifies residues amenable to mutation without compromising cellulose affinity, and informs engineering strategies for family one CBMs. PMID:26209638

  20. Structural insights into the affinity of Cel7A carbohydrate-binding module for lignin.

    PubMed

    Strobel, Kathryn L; Pfeiffer, Katherine A; Blanch, Harvey W; Clark, Douglas S

    2015-09-11

    The high cost of hydrolytic enzymes impedes the commercial production of lignocellulosic biofuels. High enzyme loadings are required in part due to their non-productive adsorption to lignin, a major component of biomass. Despite numerous studies documenting cellulase adsorption to lignin, few attempts have been made to engineer enzymes to reduce lignin binding. In this work, we used alanine-scanning mutagenesis to elucidate the structural basis for the lignin affinity of Trichoderma reesei Cel7A carbohydrate binding module (CBM). T. reesei Cel7A CBM mutants were produced with a Talaromyces emersonii Cel7A catalytic domain and screened for their binding to cellulose and lignin. Mutation of aromatic and polar residues on the planar face of the CBM greatly decreased binding to both cellulose and lignin, supporting the hypothesis that the cellulose-binding face is also responsible for lignin affinity. Cellulose and lignin affinity of the 31 mutants were highly correlated, although several mutants displayed selective reductions in lignin or cellulose affinity. Four mutants with increased cellulose selectivity (Q2A, H4A, V18A, and P30A) did not exhibit improved hydrolysis of cellulose in the presence of lignin. Further reduction in lignin affinity while maintaining a high level of cellulose affinity is thus necessary to generate an enzyme with improved hydrolysis capability. This work provides insights into the structural underpinnings of lignin affinity, identifies residues amenable to mutation without compromising cellulose affinity, and informs engineering strategies for family one CBMs. PMID:26209638

  1. A HIGH-LEVEL CALCULATION OF THE PROTON AFFINITY OF DIBORANE

    EPA Science Inventory

    The experimental proton affinity of diborane (B2H6) is based on an unstable species, B2H,+, 4 which has been observed only at low temperatures. The present work calculates the proton 5 affinity of diborane using the Gaussian-3 method and other high-level compound ab initio 6 met...

  2. Affinity for Quantitative Tools: Undergraduate Marketing Students Moving beyond Quantitative Anxiety

    ERIC Educational Resources Information Center

    Tarasi, Crina O.; Wilson, J. Holton; Puri, Cheenu; Divine, Richard L.

    2013-01-01

    Marketing students are known as less likely to have an affinity for the quantitative aspects of the marketing discipline. In this article, we study the reasons why this might be true and develop a parsimonious 20-item scale for measuring quantitative affinity in undergraduate marketing students. The scale was administered to a sample of business…

  3. Structural insights into the affinity of Cel7A carbohydrate-binding module for lignin.

    PubMed

    Strobel, Kathryn L; Pfeiffer, Katherine A; Blanch, Harvey W; Clark, Douglas S

    2015-09-11

    The high cost of hydrolytic enzymes impedes the commercial production of lignocellulosic biofuels. High enzyme loadings are required in part due to their non-productive adsorption to lignin, a major component of biomass. Despite numerous studies documenting cellulase adsorption to lignin, few attempts have been made to engineer enzymes to reduce lignin binding. In this work, we used alanine-scanning mutagenesis to elucidate the structural basis for the lignin affinity of Trichoderma reesei Cel7A carbohydrate binding module (CBM). T. reesei Cel7A CBM mutants were produced with a Talaromyces emersonii Cel7A catalytic domain and screened for their binding to cellulose and lignin. Mutation of aromatic and polar residues on the planar face of the CBM greatly decreased binding to both cellulose and lignin, supporting the hypothesis that the cellulose-binding face is also responsible for lignin affinity. Cellulose and lignin affinity of the 31 mutants were highly correlated, although several mutants displayed selective reductions in lignin or cellulose affinity. Four mutants with increased cellulose selectivity (Q2A, H4A, V18A, and P30A) did not exhibit improved hydrolysis of cellulose in the presence of lignin. Further reduction in lignin affinity while maintaining a high level of cellulose affinity is thus necessary to generate an enzyme with improved hydrolysis capability. This work provides insights into the structural underpinnings of lignin affinity, identifies residues amenable to mutation without compromising cellulose affinity, and informs engineering strategies for family one CBMs.

  4. Multiple GPCR conformations and signalling pathways: implications for antagonist affinity estimates

    PubMed Central

    Baker, Jillian G.; Hill, Stephen J.

    2007-01-01

    Antagonist affinity measurements have traditionally been considered important in characterizing the cell-surface receptors present in a particular cell or tissue. A central assumption has been that antagonist affinity is constant for a given receptor–antagonist interaction, regardless of the agonist used to stimulate that receptor or the downstream response that is measured. As a consequence, changes in antagonist affinity values have been taken as initial evidence for the presence of novel receptor subtypes. Emerging evidence suggests, however, that receptors can possess multiple binding sites and the same receptor can show different antagonist affinity measurements under distinct experimental conditions. Here, we discuss several mechanisms by which antagonists have different affinities for the same receptor as a consequence of allosterism, coupling to different G proteins, multiple (but non-interacting) receptor sites, and signal-pathway-dependent pharmacology (where the pharmacology observed varies depending on the signalling pathway measured). PMID:17629959

  5. Preparation of λN-GST fusion protein for affinity immobilization of RNA.

    PubMed

    Di Tomasso, Geneviève; Lampron, Philipe; Omichinski, James G; Legault, Pascale

    2012-01-01

    Affinity purification of in vitro transcribed RNA is becoming an attractive alternative to purification using standard denaturing gel electrophoresis. Affinity purification is particularly advantageous because it can be performed in a few hours under non-denaturing conditions. However, the performance of affinity purification methods can vary tremendously depending on the RNA immobilization matrix. It was previously shown that RNA immobilization via an optimized λN-GST fusion protein bound to glutathione-Sepharose resin allows affinity purification of RNA with very high purity and yield. This Chapter outlines the experimental procedure employed to prepare the λN-GST fusion protein used for RNA immobilization in successful affinity purifications of RNA. It describes the details of protein expression and purification as well as routine quality control analyses. PMID:23065558

  6. Low affinity binding site clusters confer hox specificity and regulatory robustness.

    PubMed

    Crocker, Justin; Abe, Namiko; Rinaldi, Lucrezia; McGregor, Alistair P; Frankel, Nicolás; Wang, Shu; Alsawadi, Ahmad; Valenti, Philippe; Plaza, Serge; Payre, François; Mann, Richard S; Stern, David L

    2015-01-15

    In animals, Hox transcription factors define regional identity in distinct anatomical domains. How Hox genes encode this specificity is a paradox, because different Hox proteins bind with high affinity in vitro to similar DNA sequences. Here, we demonstrate that the Hox protein Ultrabithorax (Ubx) in complex with its cofactor Extradenticle (Exd) bound specifically to clusters of very low affinity sites in enhancers of the shavenbaby gene of Drosophila. These low affinity sites conferred specificity for Ubx binding in vivo, but multiple clustered sites were required for robust expression when embryos developed in variable environments. Although most individual Ubx binding sites are not evolutionarily conserved, the overall enhancer architecture-clusters of low affinity binding sites-is maintained and required for enhancer function. Natural selection therefore works at the level of the enhancer, requiring a particular density of low affinity Ubx sites to confer both specific and robust expression. PMID:25557079

  7. General approach for characterizing in vitro selected peptides with protein binding affinity.

    PubMed

    Larsen, Andrew C; Gillig, Annabelle; Shah, Pankti; Sau, Sujay P; Fenton, Kathryn E; Chaput, John C

    2014-08-01

    In vitro selection technologies are important tools for identifying high affinity peptides to proteins of broad medical and biological interest. However, the technological advances that have made it possible to generate long lists of candidate peptides have far outpaced our ability to characterize the binding properties of individual peptides. Here, we describe a low cost strategy to rapidly synthesize, purify, screen, and characterize peptides for high binding affinity. Peptides are assayed in a 96-well dot blot apparatus using membranes that enable partitioning of bound and unbound peptide-protein complexes. We have validated the binding affinity constants produced by this method using known peptide ligands and applied this process to discover five new peptides with nanomolar affinity to human α-thrombin. Given the need for new analytical tools that can accelerate peptide discovery and characterization, we feel that this approach would be useful to a wide range of technologies that utilize high affinity peptides.

  8. Purification of phage display-modified bacteriophage T4 by affinity chromatography

    PubMed Central

    2011-01-01

    Background Affinity chromatography is one of the most efficient protein purification strategies. This technique comprises a one-step procedure with a purification level in the order of several thousand-fold, adaptable for various proteins, differentiated in their size, shape, charge, and other properties. The aim of this work was to verify the possibility of applying affinity chromatography in bacteriophage purification, with the perspective of therapeutic purposes. T4 is a large, icosahedral phage that may serve as an efficient display platform for foreign peptides or proteins. Here we propose a new method of T4 phage purification by affinity chromatography after its modification with affinity tags (GST and Histag) by in vivo phage display. As any permanent introduction of extraneous DNA into a phage genome is strongly unfavourable for medical purposes, integration of foreign motifs with the phage genome was not applied. The phage was propagated in bacteria expressing fusions of the phage protein Hoc with affinity tags from bacterial plasmids, independently from the phage expression system. Results Elution profiles of phages modified with the specific affinity motifs (compared to non-specific phages) document their binding to the affinity resins and effective elution with standard competitive agents. Non-specific binding was also observed, but was 102-105 times weaker than the specific one. GST-modified bacteriophages were also effectively released from glutathione Sepharose by proteolytic cleavage. The possibility of proteolytic release was designed at the stage of expression vector construction. Decrease in LPS content in phage preparations was dependent on the washing intensity; intensive washing resulted in preparations of 11-40 EU/ml. Conclusions Affinity tags can be successfully incorporated into the T4 phage capsid by the in vivo phage display technique and they strongly elevate bacteriophage affinity to a specific resin. Affinity chromatography can be

  9. Synthesis and binding affinity of an iodinated juvenile hormone

    SciTech Connect

    Prestwich, G.D.; Eng, W.S.; Robles, S.; Vogt, R.G.; Wisniewski, J.R.; Wawrzenczyk, C.

    1988-01-25

    The synthesis of the first iodinated juvenile hormone (JH) in enantiomerically enriched form is reported. This chiral compound, 12-iodo-JH I, has an iodine atom replacing a methyl group of the natural insect juvenile hormone, JH I, which is important in regulating morphogenesis and reproduction in the Lepidoptera. The unlabeled compound shows approximately 10% of the relative binding affinity for the larval hemolymph JH binding protein (JHBP) of Manduca sexta, which specifically binds natural /sup 3/H-10R,11S-JH I (labeled at 58 Ci/mmol) with a KD of 8 X 10(-8) M. It is also approximately one-tenth as biologically active as JH I in the black Manduca and epidermal commitment assays. The 12-hydroxy and 12-oxo compounds are poor competitors and are also biologically inactive. The radioiodinated (/sup 125/I)12-iodo-JH I can be prepared in low yield at greater than 2500 Ci/mmol by nucleophilic displacement using no-carrier-added /sup 125/I-labeled sodium iodide in acetone; however, synthesis using sodium iodide carrier to give the approximately 50 Ci/mmol radioiodinated ligand proceeds in higher radiochemical yield with fewer by-products and provides a radioligand which is more readily handled in binding assays. The KD of (/sup 125/I)12-iodo-JH I was determined for hemolymph JHBP of three insects: M. sexta, 795 nM; Galleria mellonella, 47 nM; Locusta migratoria, 77 nM. The selectivity of 12-iodo-JH I for the 32-kDa JHBP of M. sexta was demonstrated by direct autoradiography of a native polyacrylamide gel electrophoresis gel of larval hemolymph incubated with the radioiodinated ligand. Thus, the in vitro and in vivo activity of 12-iodo-JH I indicate that it can serve as an important new gamma-emitting probe in the search for JH receptor proteins in target tissues.

  10. Template affinity role in CH shrink by DSA planarization

    NASA Astrophysics Data System (ADS)

    Tiron, R.; Gharbi, A.; Pimenta Barros, P.; Bouanani, S.; Lapeyre, C.; Bos, S.; Fouquet, A.; Hazart, J.; Chevalier, X.; Argoud, M.; Chamiot-Maitral, G.; Barnola, S.; Monget, C.; Farys, V.; Berard-Bergery, S.; Perraud, L.; Navarro, C.; Nicolet, C.; Hadziioannou, G.; Fleury, G.

    2015-03-01

    Density multiplication and contact shrinkage of patterned templates by directed self-assembly (DSA) of block copolymers (BCP) stands out as a promising alternative to overcome the limitations of conventional lithography. The main goal of this paper is to investigate the potential of DSA to address contact and via levels patterning with high resolution by performing either CD shrink or contact multiplication. Different DSA processes are benchmarked based on several success criteria such as: CD control, defectivity (missing holes) as well as placement control. More specifically, the methodology employed to measure DSA contact overlay and the impact of process parameters on placement error control is detailed. Using the 300mm pilot line available in LETI and Arkema's materials, our approach is based on the graphoepitaxy of PS-b-PMMA block copolymers. Our integration scheme, depicted in figure 1, is based on BCP self-assembly inside organic hard mask guiding patterns obtained using 193i nm lithography. The process is monitored at different steps: the generation of guiding patterns, the directed self-assembly of block copolymers and PMMA removal, and finally the transfer of PS patterns into the metallic under layer by plasma etching. Furthermore, several process flows are investigated, either by tuning different material related parameters such as the block copolymer intrinsic period or the interaction with the guiding pattern surface (sidewall and bottom-side affinity). The final lithographic performances are finely optimized as a function of the self-assembly process parameters such as the film thickness and bake (temperature and time). Finally, DSA performances as a function of guiding patterns density are investigated. Thus, for the best integration approach, defect-free isolated and dense patterns for both contact shrink and multiplication (doubling and more) have been achieved on the same processed wafer. These results show that contact hole shrink and

  11. Affinity and its derivatives in the glass transition process.

    PubMed

    Garden, J-L; Guillou, H; Richard, J; Wondraczek, L

    2012-07-14

    The thermodynamic treatment of the glass transition remains an issue of intense debate. When associated with the formalism of non-equilibrium thermodynamics, the lattice-hole theory of liquids can provide new insight in this direction, as has been shown by Schmelzer and Gutzow [J. Chem. Phys. 125, 184511 (2006)], by Möller et al. [J. Chem. Phys. 125, 094505 (2006)], and more recently by Tropin et al. [J. Non-Cryst. Solids 357, 1291 (2011); ibid. 357, 1303 (2011)]. Here, we employ a similar approach. We include pressure as an additional variable, in order to account for the freezing-in of structural degrees of freedom upon pressure increase. Second, we demonstrate that important terms concerning first order derivatives of the affinity-driving-force with respect to temperature and pressure have been previously neglected. We show that these are of crucial importance in the approach. Macroscopic non-equilibrium thermodynamics is used to enlighten these contributions in the derivation of C(p),κ(T), and α(p). The coefficients are calculated as a function of pressure and temperature following different theoretical protocols, revealing classical aspects of vitrification and structural recovery processes. Finally, we demonstrate that a simple minimalist model such as the lattice-hole theory of liquids, when being associated with rigorous use of macroscopic non-equilibrium thermodynamics, is able to account for the primary features of the glass transition phenomenology. Notwithstanding its simplicity and its limits, this approach can be used as a very pedagogical tool to provide a physical understanding on the underlying thermodynamics which governs the glass transition process. PMID:22803545

  12. Analysis of Aged Human Serum Albumin Affinity for Doxazosin.

    PubMed

    Chudzik, Mariola; Równicka-Zubik, Joanna; Pożycka, Jadwiga; Pawelczak, Bartosz; Sulkowska, Anna

    2016-01-01

    Structural changes of human serum albumin (HSA) caused by old age and coexisting diseases result in differences in the binding of doxazosin (DOX). DOX is a postsynaptic α1- adrenoreceptor antagonist used for treatment of hypertension and benign prostatic hyperplasia. In elderly people suffering from various renal or hepatic diseases the significant portion of N-form of human serum albumin (normal) is converted to A-form (aged). The differences in binding of doxazosin to N- and Aform of albumin are an important factor, which may determines therapeutic dosage and toxicity of the test drug. To indicate these differences, the technique of fluorescence spectroscopy was used. The association constant (Ka) obtained from fluorescence quenching demonstrated that doxazosin has higher affinity for AHSA than for HSA. In order to describe the cooperativity in binding process, the values of the Hill's coefficient has been analysed. For DOX-HSA system (λex 295 nm) Hill's coefficient is close to 1 and it indicates that there is a single class of binding sites. For DOX-HSA (λex 275 nm) and DOX-AHSA (λex 275 nm and λex 295 nm) systems we observed positive cooperativity (nH>1). A greater red shift of fluorescence emission maximum of AHSA than HSA in the presence of DOX was observed. This suggests that the binding of DOX to AHSA was accompanied by a stronger increase in polarity around the fluorophores in comparison to HSA. The binding interaction between DOX and HSA has been also studied by molecular docking simulation.

  13. Determination of proton affinities and acidity constants of sugars.

    PubMed

    Feng, Shuting; Bagia, Christina; Mpourmpakis, Giannis

    2013-06-20

    Proton transfer reactions play a key role in the conversion of biomass derived sugars to chemicals. In this study, we employ high level ab initio theoretical methods, in tandem with solvation effects to calculate the proton affinities (PA) and acidity constants (pKa) of various d-glucose and d-fructose tautomers (protonation-deprotonation processes). In addition, we compare the theoretically derived pH values of sugar solutions against experimentally measured pH values in our lab. Our results demonstrate that the protonation of any of the O atoms of the sugars is thermodynamically preferred without any significant variation in the PA values. Intramolecular hydrogen transfers, dehydration reactions, and ring-opening processes were observed, resulting from the protonation of specific hydroxyl groups on the sugars. Regarding the deprotonation processes (pKa), we found that the sugars' anomeric hydroxyls exhibit the highest acidity. The theoretically calculated pH values of sugar solutions are in excellent agreement with experimental pH measurements at low sugar concentrations. At higher sugar concentrations the calculations predict less acidic solutions than the experiments. In this case, we expect the sugars to act as solvents increasing the proton solvation energy and the acidity of the solutions. We demonstrated through linear relationships that the pKa values are correlated with the relative stability of the conjugate bases. The latter is related to hydrogen bonding and polarization of the C-O(-) bond. A plausible explanation for the good performance of the direct method in calculating the pKa values of sugars can be the presence of intramolecular hydrogen bonds on the conjugate base. Both theory and experiments manifest that fructose is a stronger acid than glucose, which is of significant importance in self-catalyzed biomass-relevant dehydration reactions. PMID:23706015

  14. High-affinity binding of fibronectin to cultured Kupffer cells

    SciTech Connect

    Cardarelli, P.M.; Blumenstock, F.A.; McKeown-Longo, P.J.; Saba, T.M.; Mazurkiewicz, J.E.; Dias, J.A. )

    1990-11-01

    Hepatic Kupffer cells are a major component of the reticuloendothelial or macrophage system. They were the first phagocytic cell type whose phagocytosis was shown to be influenced by plasma fibronectin, a dimeric opsonic glycoprotein. In the current study, the binding of soluble radioiodinated fibronectin purified from rat serum to isolated rat hepatic Kupffer cells was investigated using a cultured Kupffer cell monolayer technique. Binding was specific, since unlabeled purified fibronectin competed in a dose-dependent manner with the 125I-fibronectin for binding to the Kupffer cells. Addition of gelatin enhanced the binding of 125I-fibronectin to Kupffer cells. The phagocytosis of gelatinized-coated red cells by Kupffer cells was increased either by preopsonizing the target particles with purified fibronectin or by the addition of purified fibronectin to the culture medium. In contrast, exposure of the Kupffer cells to medium containing purified fibronectin followed by wash-removal of the fibronectin did not increase the uptake of gelatin-coated red blood cells, even though fibronectin was detected on the surface of the Kupffer cells by immunofluorescence. Trypsinized monolayers expressed decreased capacity to bind 125I-fibronectin as well as fibronectin-coated sheep erythrocytes. The binding of 125I-fibronectin-gelatin complexes was inhibited by excess unlabeled fibronectin. We calculated that specific high-affinity (Kd = 7.46 x 10(-9) M) binding sites for fibronectin exist on Kupffer cells. There are approximately 2,800-3,500 binding sites or putative fibronectin receptors per Kupffer cell. These sites appear to mediate the enhanced phagocytosis of gelatin-coated particles opsonized by fibronectin.

  15. Binding Kinetics versus Affinities in BRD4 Inhibition.

    PubMed

    Kuang, Ming; Zhou, Jingwei; Wang, Laiyou; Liu, Zhihong; Guo, Jiao; Wu, Ruibo

    2015-09-28

    Bromodomains (BRDs) are protein modules that selectively recognize histones as a "reader" by binding to an acetylated lysine substrate. The human BRD4 has emerged as a promising drug target for a number of disease pathways, and several potent BRD inhibitors have been discovered experimentally recently. However, the detailed inhibition mechanism especially for the inhibitor binding kinetics is not clear. Herein, by employing classical molecular dynamics (MD) and state-of-the-art density functional QM/MM MD simulations, the dynamic characteristics of ZA-loop in BRD4 are revealed. And then the correlation between binding pocket size and ZA-loop motion is elucidated. Moreover, our simulations found that the compound (-)-JQ1 could be accommodated reasonably in thermodynamics whereas it is infeasible in binding kinetics against BRD4. Its racemate (+)-JQ1 proved to be both thermodynamically reasonable and kinetically achievable against BRD4, which could explain the previous experimental results that (+)-JQ1 shows a high inhibitory effect toward BRD4 (IC50 is 77 nM) while (-)-JQ1 is inactive (>10 μM). Furthermore, the L92/L94/Y97 in the ZA-loop and Asn140 in the BC-loop are identified to be critical residues in (+)-JQ1 binding/releasing kinetics. All these findings shed light on further selective inhibitor design toward BRD family, by exploiting the non-negligible ligand binding kinetics features and flexible ZA-loop motions of BRD, instead of only the static ligand-protein binding affinity. PMID:26263125

  16. Genetic Affinities of the Central Indian Tribal Populations

    PubMed Central

    Singh, Vipin Kumar; Shah, Anish M.; Anugula, Sharath; Rani, Deepa Selvi; Reddy, Alla G.; Eaaswarkhanth, Muthukrishnan; Chaubey, Gyaneshwer; Singh, Lalji; Thangaraj, Kumarasamy

    2012-01-01

    Background The central Indian state Madhya Pradesh is often called as ‘heart of India’ and has always been an important region functioning as a trinexus belt for three major language families (Indo-European, Dravidian and Austroasiatic). There are less detailed genetic studies on the populations inhabited in this region. Therefore, this study is an attempt for extensive characterization of genetic ancestries of three tribal populations, namely; Bharia, Bhil and Sahariya, inhabiting this region using haploid and diploid DNA markers. Methodology/Principal Findings Mitochondrial DNA analysis showed high diversity, including some of the older sublineages of M haplogroup and prominent R lineages in all the three tribes. Y-chromosomal biallelic markers revealed high frequency of Austroasiatic-specific M95-O2a haplogroup in Bharia and Sahariya, M82-H1a in Bhil and M17-R1a in Bhil and Sahariya. The results obtained by haploid as well as diploid genetic markers revealed strong genetic affinity of Bharia (a Dravidian speaking tribe) with the Austroasiatic (Munda) group. The gene flow from Austroasiatic group is further confirmed by their Y-STRs haplotype sharing analysis, where we determined their founder haplotype from the North Munda speaking tribe, while, autosomal analysis was largely in concordant with the haploid DNA results. Conclusions/Significance Bhil exhibited largely Indo-European specific ancestry, while Sahariya and Bharia showed admixed genetic package of Indo-European and Austroasiatic populations. Hence, in a landscape like India, linguistic label doesn't unequivocally follow the genetic footprints. PMID:22393414

  17. Binding Kinetics versus Affinities in BRD4 Inhibition.

    PubMed

    Kuang, Ming; Zhou, Jingwei; Wang, Laiyou; Liu, Zhihong; Guo, Jiao; Wu, Ruibo

    2015-09-28

    Bromodomains (BRDs) are protein modules that selectively recognize histones as a "reader" by binding to an acetylated lysine substrate. The human BRD4 has emerged as a promising drug target for a number of disease pathways, and several potent BRD inhibitors have been discovered experimentally recently. However, the detailed inhibition mechanism especially for the inhibitor binding kinetics is not clear. Herein, by employing classical molecular dynamics (MD) and state-of-the-art density functional QM/MM MD simulations, the dynamic characteristics of ZA-loop in BRD4 are revealed. And then the correlation between binding pocket size and ZA-loop motion is elucidated. Moreover, our simulations found that the compound (-)-JQ1 could be accommodated reasonably in thermodynamics whereas it is infeasible in binding kinetics against BRD4. Its racemate (+)-JQ1 proved to be both thermodynamically reasonable and kinetically achievable against BRD4, which could explain the previous experimental results that (+)-JQ1 shows a high inhibitory effect toward BRD4 (IC50 is 77 nM) while (-)-JQ1 is inactive (>10 μM). Furthermore, the L92/L94/Y97 in the ZA-loop and Asn140 in the BC-loop are identified to be critical residues in (+)-JQ1 binding/releasing kinetics. All these findings shed light on further selective inhibitor design toward BRD family, by exploiting the non-negligible ligand binding kinetics features and flexible ZA-loop motions of BRD, instead of only the static ligand-protein binding affinity.

  18. A global/local affinity graph for image segmentation.

    PubMed

    Xiaofang Wang; Yuxing Tang; Masnou, Simon; Liming Chen

    2015-04-01

    Construction of a reliable graph capturing perceptual grouping cues of an image is fundamental for graph-cut based image segmentation methods. In this paper, we propose a novel sparse global/local affinity graph over superpixels of an input image to capture both short- and long-range grouping cues, and thereby enabling perceptual grouping laws, including proximity, similarity, continuity, and to enter in action through a suitable graph-cut algorithm. Moreover, we also evaluate three major visual features, namely, color, texture, and shape, for their effectiveness in perceptual segmentation and propose a simple graph fusion scheme to implement some recent findings from psychophysics, which suggest combining these visual features with different emphases for perceptual grouping. In particular, an input image is first oversegmented into superpixels at different scales. We postulate a gravitation law based on empirical observations and divide superpixels adaptively into small-, medium-, and large-sized sets. Global grouping is achieved using medium-sized superpixels through a sparse representation of superpixels' features by solving a ℓ0-minimization problem, and thereby enabling continuity or propagation of local smoothness over long-range connections. Small- and large-sized superpixels are then used to achieve local smoothness through an adjacent graph in a given feature space, and thus implementing perceptual laws, for example, similarity and proximity. Finally, a bipartite graph is also introduced to enable propagation of grouping cues between superpixels of different scales. Extensive experiments are carried out on the Berkeley segmentation database in comparison with several state-of-the-art graph constructions. The results show the effectiveness of the proposed approach, which outperforms state-of-the-art graphs using four different objective criteria, namely, the probabilistic rand index, the variation of information, the global consistency error, and the

  19. A global/local affinity graph for image segmentation.

    PubMed

    Xiaofang Wang; Yuxing Tang; Masnou, Simon; Liming Chen

    2015-04-01

    Construction of a reliable graph capturing perceptual grouping cues of an image is fundamental for graph-cut based image segmentation methods. In this paper, we propose a novel sparse global/local affinity graph over superpixels of an input image to capture both short- and long-range grouping cues, and thereby enabling perceptual grouping laws, including proximity, similarity, continuity, and to enter in action through a suitable graph-cut algorithm. Moreover, we also evaluate three major visual features, namely, color, texture, and shape, for their effectiveness in perceptual segmentation and propose a simple graph fusion scheme to implement some recent findings from psychophysics, which suggest combining these visual features with different emphases for perceptual grouping. In particular, an input image is first oversegmented into superpixels at different scales. We postulate a gravitation law based on empirical observations and divide superpixels adaptively into small-, medium-, and large-sized sets. Global grouping is achieved using medium-sized superpixels through a sparse representation of superpixels' features by solving a ℓ0-minimization problem, and thereby enabling continuity or propagation of local smoothness over long-range connections. Small- and large-sized superpixels are then used to achieve local smoothness through an adjacent graph in a given feature space, and thus implementing perceptual laws, for example, similarity and proximity. Finally, a bipartite graph is also introduced to enable propagation of grouping cues between superpixels of different scales. Extensive experiments are carried out on the Berkeley segmentation database in comparison with several state-of-the-art graph constructions. The results show the effectiveness of the proposed approach, which outperforms state-of-the-art graphs using four different objective criteria, namely, the probabilistic rand index, the variation of information, the global consistency error, and the

  20. Affinity Interaction between Hexamer Peptide Ligand HWRGWV and Immunoglobulin G Studied by Quartz Crystal Microbalance and Surface Plasmon Resonance

    NASA Astrophysics Data System (ADS)

    Shen, Fei

    Immunoglobulins (Ig), also referred to as antibodies, act as protective agents against pathogens trying to invade an organism. Human immunoglobulin G (hIgG), as the most prominent immunoglobulin presented in serum and other human fluids, has broad applications in fields like immunotherapy and clinical diagnostics. Staphylococcus aureus Protein A and Streptococcus Protein G are the most common affinity ligands for IgG purifaction and detection. However, drawbacks associated with these two protein ligands have motivated searches for alternative affinity ligands. The hexamer peptide ligand HWRGWV identified from a one-bead-one-peptide combinatorial library synthesized on chromatography resins has demonstrated high affinity and specificity to the Fc fragment of hIgG. A chromatography resin with HWRGWV can purify human IgG (hIgG) from complete minimum essential medium (cMEM) with purities and yields as high as 95%, which are comparable to using Protein A as affinity ligand (4). As a short peptide ligand, HWRGWV can be produced at relatively low costs under good manufacturing practices (GMP) conditions, it is highly robust, less immunogenic and allows for milder elution conditions for the bound antibody (3, 5). Although this short peptide ligand has exhibited promising properties for IgG capture and purification, limited information is available on the intrinsic mechanisms of affinity interaction between the peptide ligand and target protein. In this study, the affinity interaction between hIgG and peptide ligand immobilized on solid surfaces was studied by quartz crystal microbalance (QCM) and surface plasmon resonance (SPR). Compared with previous methods employed for the peptide characterization, QCM and SPR can provide direct measurements of equilibrium adsorption isotherms and rates of adsorption, allowing a complete kinetic and thermodynamics analyses of the ligand-target interactions. New methods were developed to modify gold and silica surfaces of QCM and SPR

  1. Traces of Intertwiners for Quantum Affine sl_2 and Felder-Varchenko Functions

    NASA Astrophysics Data System (ADS)

    Sun, Yi

    2016-10-01

    We show that the traces of {U_q({widehat{sl}_2)}-intertwiners of [ESV02] valued in the three-dimensional evaluation representation converge in a certain region of parameters and give a representation-theoretic construction of Felder-Varchenko's hypergeometric solutions to the q-KZB heat equation given in [FV02]. This gives the first proof that such a trace function converges and resolves the first case of the Etingof-Varchenko conjecture of [EV00]. As applications, we prove a symmetry property for traces of intertwiners and prove Felder-Varchenko's conjecture in [FV04] that their elliptic Macdonald polynomials are related to the affine Macdonald polynomials defined as traces over irreducible integrable {U_q({widehat{sl}_2)}-modules in [EK95]. In the trigonometric and classical limits, we recover results of [EK94,EV00]. Our method relies on an interplay between the method of coherent states applied to the free field realization of the q-Wakimoto module of [Mat94], convergence properties given by the theta hypergeometric integrals of [FV02], and rationality properties originating from the representation-theoretic definition of the trace function.

  2. [Ecological affinity and current distribution of primates (Cebidae) in Campeche, Mexico].

    PubMed

    Navarro Fernández, Eloísa; Pozo de la Tijera, Carmen; Escobedo Cabrera, Enrique

    2003-06-01

    We carried out surveys realized field work from March to September 2000 to get the current distribution of Cebids in the state of Campeche, Mexico. Based on interviews and direct observations. We defined the distribution of Ateles geoffroyi yucatanensis and Alouatta pigra and we documented the first time localities where Allouata palliata is found in the state. We made distributional maps of each species using vegetation overlays from Inventario Nacional Forestal (Inv For) and each point documented during fieldwork. We presented the distribution of species according to confiability of the verified or expected data. Using the attributes table of Inv For, we calculated the areas of distribution which were 22,735 km2 for Alouatta sp. and 18,501 km2 for A. g. yucatanensis. We also presented the area occupied by each species according to vegetation types and the relative proportion of these vegetation types in the state. We confirmed the ability of Alouatta sp. to survive in disturbed environments produced by habitat fragmentation, and the affinity of A. g. yucatanesis to well preserved habitats.

  3. Physics-based enzyme design: predicting binding affinity and catalytic activity.

    PubMed

    Sirin, Sarah; Pearlman, David A; Sherman, Woody

    2014-12-01

    Computational enzyme design is an emerging field that has yielded promising success stories, but where numerous challenges remain. Accurate methods to rapidly evaluate possible enzyme design variants could provide significant value when combined with experimental efforts by reducing the number of variants needed to be synthesized and speeding the time to reach the desired endpoint of the design. To that end, extending our computational methods to model the fundamental physical-chemical principles that regulate activity in a protocol that is automated and accessible to a broad population of enzyme design researchers is essential. Here, we apply a physics-based implicit solvent MM-GBSA scoring approach to enzyme design and benchmark the computational predictions against experimentally determined activities. Specifically, we evaluate the ability of MM-GBSA to predict changes in affinity for a steroid binder protein, catalytic turnover for a Kemp eliminase, and catalytic activity for α-Gliadin peptidase variants. Using the enzyme design framework developed here, we accurately rank the most experimentally active enzyme variants, suggesting that this approach could provide enrichment of active variants in real-world enzyme design applications.

  4. The Lectin Frontier Database (LfDB), and data generation based on frontal affinity chromatography.

    PubMed

    Hirabayashi, Jun; Tateno, Hiroaki; Shikanai, Toshihide; Aoki-Kinoshita, Kiyoko F; Narimatsu, Hisashi

    2015-01-01

    Lectins are a large group of carbohydrate-binding proteins, having been shown to comprise at least 48 protein scaffolds or protein family entries. They occur ubiquitously in living organisms-from humans to microorganisms, including viruses-and while their functions are yet to be fully elucidated, their main underlying actions are thought to mediate cell-cell and cell-glycoconjugate interactions, which play important roles in an extensive range of biological processes. The basic feature of each lectin's function resides in its specific sugar-binding properties. In this regard, it is beneficial for researchers to have access to fundamental information about the detailed oligosaccharide specificities of diverse lectins. In this review, the authors describe a publicly available lectin database named "Lectin frontier DataBase (LfDB)", which undertakes the continuous publication and updating of comprehensive data for lectin-standard oligosaccharide interactions in terms of dissociation constants (Kd's). For Kd determination, an advanced system of frontal affinity chromatography (FAC) is used, with which quantitative datasets of interactions between immobilized lectins and >100 fluorescently labeled standard glycans have been generated. The FAC system is unique in its clear principle, simple procedure and high sensitivity, with an increasing number (>67) of associated publications that attest to its reliability. Thus, LfDB, is expected to play an essential role in lectin research, not only in basic but also in applied fields of glycoscience. PMID:25580689

  5. 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.

  6. Flow of power-law fluids in self-affine fracture channels.

    PubMed

    Yan, Yiguang; Koplik, Joel

    2008-03-01

    The two-dimensional pressure driven flow of non-Newtonian power-law fluids in self-affine fracture channels at finite Reynolds number is calculated. The channels have constant mean aperture and two values zeta=0.5 and 0.8 of the Hurst exponent are considered. The calculation is based on the lattice-Boltzmann method, using a different technique to obtain a power-law variation in viscosity, and the behavior of shear-thinning, Newtonian, and shear-thickening liquids is compared. Local aspects of the flow fields, such as maximum velocity and pressure fluctuations, are studied, and the non-Newtonian fluids are compared to the (previously studied) Newtonian case. We find a scaling relation between permeability and mean aperture in the low Reynolds number regime, generalizing an earlier result for Newtonian fluids. As the Reynolds number increases, we observe the same sequence of transitions to nonlinearity found in intergranular porous media. Furthermore, the permeability results may be collapsed into a master curve of friction factor vs Reynolds number, using a scaling similar to that employed for power-law fluids in porous media.

  7. Analysis of affinities between specific biological ligands using atomic force microscopy.

    PubMed

    Hu, Xiao; Dinu, Cerasela Zoica

    2015-12-21

    In the cell, protein-ligand recognition involves association and dissociation processes controlled by the affinity of the two binding partners and chemical harvesting of adenosine triphosphate energy. Fundamental knowledge of selected recognition events is currently translated in a synthetic environment for biosensors, immunoassays and diagnosis applications, or for pharmaceutical development. However, in order to advance such fields, one needs to determine the lifetime and binding efficiency of the two partners, as well as the complex energy landscape parameters. We employed contact mode atomic force microscopy to evaluate the association and dissociation events between streptavidin protein and its anti-streptavidin antibody ligand currently used for nucleotide array, ELISA, and flow cytometry applications, just to name a few. Using biotin as the control, our analysis helped characterize and differentiate multi- or single bonds of different strengths as well as associated energy landscapes to determine the protein-ligand structural arrangement at nanointerfaces and how these depend on the specificity of the ligand-recognition reaction. Our results suggest that understanding the importance of the rupture forces between a protein and its ligand could serve as the first step to protect on-off switches for biomedical research applications where specificity and selectivity are foremost sought. PMID:26525901

  8. Maturation of functional antibody affinity in animals immunised with synthetic foot-and-mouth disease virus.

    PubMed

    Mulcahy, G; Reid, E; Dimarchi, R D; Gale, C; Doel, T R

    1992-03-01

    A good correlation exists between specific neutralising antibody titre and protection against challenge with foot-and-mouth disease virus (FMDV) in infected or virus-vaccinated cattle, but not in the case of animals immunised with synthetic FMDV peptides. Therefore, mechanisms other than simple neutralisation are likely to be important in vivo. Antibody affinity may influence the protective capacity of sera from immunised animals and experiments were carried out to measure the functional affinity for synthetic FMDV peptide of sera from guinea pigs and cattle given various synthetic vaccines. In guinea pigs given a single dose of synthetic vaccine, antibody affinity increased with time after immunisation. In cattle, however, administration of a second dose of peptide 21 days after the first markedly retarded the process of affinity maturation. For guinea pig sera of equivalent neutralising activity, those of higher functional affinity had higher protective indices than those of lower functional affinity. Knowledge of the importance of antibody affinity in protection against FMD is important for an improved understanding of the mechanisms of protection and for the design of novel vaccines. PMID:1316628

  9. Specificity and Affinity Quantification of Flexible Recognition from Underlying Energy Landscape Topography

    PubMed Central

    Chu, Xiakun; Wang, Jin

    2014-01-01

    Flexibility in biomolecular recognition is essential and critical for many cellular activities. Flexible recognition often leads to moderate affinity but high specificity, in contradiction with the conventional wisdom that high affinity and high specificity are coupled. Furthermore, quantitative understanding of the role of flexibility in biomolecular recognition is still challenging. Here, we meet the challenge by quantifying the intrinsic biomolecular recognition energy landscapes with and without flexibility through the underlying density of states. We quantified the thermodynamic intrinsic specificity by the topography of the intrinsic binding energy landscape and the kinetic specificity by association rate. We found that the thermodynamic and kinetic specificity are strongly correlated. Furthermore, we found that flexibility decreases binding affinity on one hand, but increases binding specificity on the other hand, and the decreasing or increasing proportion of affinity and specificity are strongly correlated with the degree of flexibility. This shows more (less) flexibility leads to weaker (stronger) coupling between affinity and specificity. Our work provides a theoretical foundation and quantitative explanation of the previous qualitative studies on the relationship among flexibility, affinity and specificity. In addition, we found that the folding energy landscapes are more funneled with binding, indicating that binding helps folding during the recognition. Finally, we demonstrated that the whole binding-folding energy landscapes can be integrated by the rigid binding and isolated folding energy landscapes under weak flexibility. Our results provide a novel way to quantify the affinity and specificity in flexible biomolecular recognition. PMID:25144525

  10. Characterization of opiate receptor heterogeneity using affinity ligands and phospholipase A/sub 2/

    SciTech Connect

    Reichman, M.

    1985-01-01

    The primary aim of the dissertation was to study the heterogeneity of opiate receptors by utilizing affinity ligands, and by modification of the receptor lipid-microenvironment with phospholipase A/sub 2/ (PLA/sub 2/). The affinity ligands, 14-bromacetamidomorphine (BAM) and 14-chloroacetylnaltrexone (CAN), selectively inactivated high affinity dihydromorphine binding sites in an apparently irreversible manner (the inhibition was resistant to extensive washes of treated neural membrane homogenates). The inhibitory effect of PLA/sub 2/ (10 ng/ml) on opiate receptor subtypes was determined using (/sup 3/H)-dihydromorphine (..mu..-type agonist), (/sup 3/H)-enkephalin (delta agonist) and (/sup 3/H)-naloxone (..mu.. antagonist). PLA/sub 2/ abolished the high affinity antagonist binding site, whereas it inhibited high and low affinity agonist binding sites similarly. The results suggest that high affinity antagonist binding sites are different from high affinity agonist binding sites. Indirect binding assays demonstrated that the selectivities of ..mu..- and delta receptors are not affected significantly by PLA/sub 2/ treatment.

  11. Computational protein design suggests that human PCNA-partner interactions are not optimized for affinity.

    PubMed

    Fridman, Yearit; Gur, Eyal; Fleishman, Sarel J; Aharoni, Amir

    2013-02-01

    Increasing the affinity of binding proteins is invaluable for basic and applied biological research. Currently, directed protein evolution experiments are the main approach for generating such proteins through the construction and screening of large mutant libraries. Proliferating cell nuclear antigen (PCNA) is an essential hub protein that interacts with many different partners to tightly regulate DNA replication and repair in all eukaryotes. Here, we used computational design to generate human PCNA mutants with enhanced affinity for several different partners. We identified double mutations in PCNA, outside the main partner binding site, that were predicted to increase PCNA-partner binding affinities compared to the wild-type protein by forming additional hydrophobic interactions with conserved residues in the PCNA partners. Affinity increases were experimentally validated with four different PCNA partners, demonstrating that computational design can reveal unexpected regions where affinity enhancements in natural systems are possible. The designed PCNA mutants can be used as a valuable tool for further examination of the regulation of PCNA-partner interactions during DNA replication and repair both in vitro and in vivo. More broadly, the ability to engineer affinity increases toward several PCNA partners suggests that interaction affinity is not an evolutionarily optimized trait of this system. PMID:23011891

  12. Proton Affinity of Isomeric Dipeptides Containing Lysine and Non-Proteinogenic Lysine Homologues.

    PubMed

    Batoon, Patrick; Ren, Jianhua

    2016-08-18

    Conformational effects on the proton affinity of oligopeptides have been studied using six alanine (A)-based acetylated dipeptides containing a basic probe that is placed closest to either the C- or the N-terminus. The basic probe includes Lysine (Lys) and two nonproteinogenic Lys-homologues, ornithine (Orn) and 2,3-diaminopropionic acid (Dap). The proton affinities of the peptides have been determined using the extended Cooks kinetic method in a triple quadrupole mass spectrometer. Computational studies have been carried out to search for the lowest energy conformers and to calculate theoretical proton affinities as well as various molecular properties using the density functional theory. The dipeptides containing a C-terminal probe, ALys, AOrn, and ADap, were determined to have a higher proton affinity by 1-4 kcal/mol than the corresponding dipeptides containing an N-terminal probe, LysA, OrnA, and DapA. For either the C-probe peptides or the N-probe peptides, the proton affinity reduces systematically as the side-chain of the probe residue is shortened. The difference in the proton affinities between isomeric peptides is largely associated with the variation of the conformations. The peptides with higher values of the proton affinity adopt a relatively compact conformation such that the protonated peptides can be stabilized through more efficient internal solvation. PMID:27459294

  13. Affinity chromatography of nicotinamide-adenine dinucleotide-linked dehydrogenases on immobilized derivatives of the dinucleotide.

    PubMed

    Barry, S; O'Carra, P

    1973-12-01

    1. Three established methods for immobilization of ligands through primary amino groups promoted little or no attachment of NAD(+) through the 6-amino group of the adenine residue. Two of these methods (coupling to CNBr-activated agarose and to carbodi-imide-activated carboxylated agarose derivatives) resulted instead in attachment predominantly through the ribosyl residues. Other immobilized derivatives were prepared by azolinkage of NAD(+) (probably through the 8 position of the adenine residue) to a number of different spacer-arm-agarose derivatives. 2. The effectiveness of these derivatives in the affinity chromatography of a variety of NAD-linked dehydrogenases was investigated, applying rigorous criteria to distinguish general or non-specific adsorption effects from truly NAD-specific affinity (bio-affinity). The ribosyl-attached NAD(+) derivatives displayed negligible bio-affinity for any of the NAD-linked dehydrogenases tested. The most effective azo-linked derivative displayed strong bio-affinity for glycer-aldehyde 3-phosphate dehydrogenase, weaker bio-affinity for lactate dehydrogenase and none at all for malate dehydrogenase, although these three enzymes have very similar affinities for soluble NAD(+). Alcohol dehydrogenase and xanthine dehydrogenase were subject to such strong non-specific interactions with the hydrocarbon spacer-arm assembly that any specific affinity was completely eclipsed. 3. It is concluded that, in practice, the general effectiveness of a general ligand may be considerably distorted and attenuated by the nature of the immobilization linkage. However, this attenuation can result in an increase in specific effectiveness, allowing dehydrogenases to be separated from one another in a manner unlikely to be feasible if the general effectiveness of the ligand remained intact. 4. The bio-affinity of the various derivatives for lactate dehydrogenase is correlated with the known structure of the NAD(+)-binding site of this enzyme. Problems

  14. Anion-induced increases in the affinity of colcemid binding to tubulin.

    PubMed

    Ray, K; Bhattacharyya, B; Biswas, B B

    1984-08-01

    Colcemid binds tubulin rapidly and reversibly in contrast to colchicine which binds tubulin relatively slowly and essentially irreversibly. At 37 degrees C the association rate constant for colcemid binding is 1.88 X 10(6) M-1 h-1, about 10 times higher than that for colchicine; this is reflected in the activation energies for binding which are 51.4 kJ/mol for colcemid and 84.8 kJ/mol for colchicine. Scatchard analysis indicates two binding sites on tubulin having different affinities for colcemid. The high-affinity site (Ka = 0.7 X 10(5) M-1 at 37 degrees C) is sensitive to temperature and binds both colchicine and colcemid and hence they are mutually competitive inhibitors. The low-affinity site (Kb = 1.2 X 10(4) M-1) is rather insensitive to temperature and binds only colcemid. Like colchicine, 0.6 mol of colcemid are bound/mol of tubulin dimer (at the high-affinity site) and the reaction is entropy driven (163 J K-1 mol-1). Similar to colchicine, colcemid binding to tubulin is stimulated by certain anions (viz. sulfate and tartrate) but by a different mechanism. Colcemid binding affinity at the lower-affinity site of tubulin is increased in the presence of ammonium sulfate. Interestingly, the lower-affinity site on tubulin for colcemid, even when converted to higher affinity in presence of ammonium sulfate, is not recognized by colchicine. We conclude that tubulin possesses two binding sites, one of which specifically recognized the groups present on the B-ring of colchicine molecule and is effected by the ammonium sulfate, whereas the higher-affinity site, which could accommodate both colchicine and colcemid, possibly recognized the A and C ring of colchicine.

  15. Methyl cation affinities of neutral and anionic maingroup-element hydrides: trends across the periodic table and correlation with proton affinities.

    PubMed

    Mulder, R Joshua; Guerra, Célia Fonseca; Bickelhaupt, F Matthias

    2010-07-22

    We have computed the methyl cation affinities in the gas phase of archetypal anionic and neutral bases across the periodic table using ZORA-relativistic density functional theory (DFT) at BP86/QZ4P//BP86/TZ2P. The main purpose of this work is to provide the methyl cation affinities (and corresponding entropies) at 298 K of all anionic (XH(n-1)(-)) and neutral bases (XH(n)) constituted by maingroup-element hydrides of groups 14-17 and the noble gases (i.e., group 18) along the periods 2-6. The cation affinity of the bases decreases from H(+) to CH(3)(+). To understand this trend, we have carried out quantitative bond energy decomposition analyses (EDA). Quantitative correlations are established between the MCA and PA values.

  16. Methyl cation affinities of neutral and anionic maingroup-element hydrides: trends across the periodic table and correlation with proton affinities.

    PubMed

    Mulder, R Joshua; Guerra, Célia Fonseca; Bickelhaupt, F Matthias

    2010-07-22

    We have computed the methyl cation affinities in the gas phase of archetypal anionic and neutral bases across the periodic table using ZORA-relativistic density functional theory (DFT) at BP86/QZ4P//BP86/TZ2P. The main purpose of this work is to provide the methyl cation affinities (and corresponding entropies) at 298 K of all anionic (XH(n-1)(-)) and neutral bases (XH(n)) constituted by maingroup-element hydrides of groups 14-17 and the noble gases (i.e., group 18) along the periods 2-6. The cation affinity of the bases decreases from H(+) to CH(3)(+). To understand this trend, we have carried out quantitative bond energy decomposition analyses (EDA). Quantitative correlations are established between the MCA and PA values. PMID:20575582

  17. Indium phosphide negative electron affinity photocathodes: Surface cleaning and activation

    NASA Astrophysics Data System (ADS)

    Sun, Yun

    InP(100) is a very important semi-conductor for many applications. When activated by Cs and oxygen, the InP surface achieves the state of Negative Electron Affinity (NEA) making the Cs+O/InP system a very efficient electron source. Despite many years of study, the chemical cleaning and activation of InP are still not well understood. In our work, we have established an understanding of the basic physics and chemistry for the chemical cleaning and activation of the InP(100) surface. Synchrotron Radiation Photoelectron Spectroscopy is the main technique used in this study because of its high surface sensitivity and ability to identify chemical species present on the surface at each stage of our process. A clean, stoichiometric InP(100) surface is crucial for obtaining high performance of NEA photocathodes. Therefore, the first part of our study focused on the chemical cleaning of InP(100). We found that hydrogen peroxide based solutions alone, originally developed to clean GaAs(100) surfaces and widely used for InP(100), do not result in clean InP(I00) surfaces because oxide is left on the surface. A second cleaning step, which uses acid solutions like HCl or H2SO4, can remove all the oxide and leave a 0.4 ML protective layer of elemental phosphorous on the surface. The elemental phosphorous can be removed by annealing at 330°C and a clean InP(100) surface can be obtained. Cs deposition on InP(100) surface shows clear charge transfer from the Cs ad-atoms to the substrate. When the Cs/InP(100) surface is dosed with oxygen, the charge transfer from the Cs to substrate is reduced and substrate is oxidized. The activation of InP as a NEA photocathode is carried out by an alternating series of steps consisting of Cs deposition and Cs+O co-deposition. Two types of oxygen are found after activation. The first is dissociated oxygen and the other is a di-oxygen species (peroxide or superoxide). The decay of quantum-yield with time and with annealing is studied and changes in

  18. (/sup 14/C)chloroacetylcholine as an advantageous affinity label of the acetylcholine receptor

    SciTech Connect

    Bodmer, D.M.; Sin-Ren, A.C.; Waser, P.G.

    1987-01-01

    The alkylating agent (/sup 14/C)chloroacetylcholine perchlorate ((/sup 14/C) ClACh) was synthesized and used for affinity labelling of the nicotinic acetylcholine receptor from Torpedo marmorata. Solubilized and affinity-purified receptor proteins were reduced and alkylated according to the bromoacetylcholine-method. Covalent binding of (/sup 14/C) ClACh to the cholinergic receptor proved to be specific and saturable, and occurred exclusively to the alpha-subunit. Halogen substitution of acetylcholine by chlorine and insertion of a /sup 14/C-isotope instead of the widely used /sup 3/H resulted in favorable properties of the affinity label.

  19. 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.

  20. Use of Tandem Affinity Chromatography for Purification of Cannabinoid Receptor CB2

    PubMed Central

    Locatelli-Hoops, Silvia C.; Yeliseev, Alexei A.

    2016-01-01

    Tandem affinity purification has been increasingly applied to isolation of recombinant proteins. It relies on two consecutive chromatographic steps that take advantage of the affinity tags placed at opposing ends of the target protein. This allows for efficient removal of contaminating proteins, including products of proteolytic degradation of the fusion that lack either N- or C-terminal tags. Here, we describe the use of two small affinity tags, a poly-histidine tag and a Strep-tag for expression and purification of the human cannabinoid receptor CB2, an integral membrane G protein-coupled receptor. PMID:24943318

  1. Affinity of pyridylalkylamines for nicotinic, muscarinic and histaminic recognition sites in brain tissue preparations.

    PubMed

    Repond, C; Pratt, J A; Stolerman, I P; Mayer, J M; Jenner, P; Marsden, C D; Testa, B

    1986-08-01

    The affinity of 15 regioisomeric and homologous pyridylalkylamines was examined in brain preparations for nicotinic, muscarinic, and H1-histaminic binding sites as labeled by [3H]-nicotine, [3H]-dexetimide and [3H]-mepyramine, respectively. Overall, the compounds show a clear selectivity for the nicotinic versus muscarinic binding sites, and a weak affinity for the H1-histaminic sites. Variations in affinity appear to be partly influenced by steric factors (such as position of attachment, length and rigidity of side-chain) and marginally by lipophilicity. PMID:3778556

  2. Symmetries of Type N Pure Radiation Fields

    NASA Astrophysics Data System (ADS)

    Ahsan, Zafar; Ali, Musavvir

    2015-05-01

    The geometrical symmetries corresponding to the continuous groups of collineations and motions generated by a null vector l are considered. These symmetries have been translated into the language of Newman-Penrose formalism for pure radiation (PR) type N fields. It is seen that for such fields, conformal, special conformal and homothetic motions degenerate to motion. The concept of free curvature, matter curvature and matter affine collineations have been discussed and the conditions under which PR type N fields admit such collineations have been obtained. Moreover, it is shown that the projective collineation degenerate to matter affine, special projective, conformal, special conformal, null geodesic and special null geodesic collineations. It is also seen that type N pure radiation fields admit Maxwell collineation along the propagation vector l.

  3. Affinity Chromatography of Native and Recombinant Proteins from Receptors for Insulin and IGF-I to Recombinant Single Chain Antibodies.

    PubMed

    Fujita-Yamaguchi, Yoko

    2015-01-01

    Affinity chromatography is an efficient method to isolate proteins by taking advantage of their affinities for specific molecules such as substrates, inhibitors, antigens, ligands, antibodies, and other interacting molecules, including subunits. Nowadays, we take the effectiveness and excellence of this technology for granted. This essay will mainly cover the use of affinity chromatography based on my experience. PMID:26579073

  4. Affinity Chromatography of Native and Recombinant Proteins from Receptors for Insulin and IGF-I to Recombinant Single Chain Antibodies

    PubMed Central

    Fujita-Yamaguchi, Yoko

    2015-01-01

    Affinity chromatography is an efficient method to isolate proteins by taking advantage of their affinities for specific molecules such as substrates, inhibitors, antigens, ligands, antibodies, and other interacting molecules, including subunits. Nowadays, we take the effectiveness and excellence of this technology for granted. This essay will mainly cover the use of affinity chromatography based on my experience. PMID:26579073

  5. Tuned-Affinity Bivalent Ligands for the Characterization of Opioid Receptor Heteromers.

    PubMed

    Harvey, Jessica H; Long, Darcie H; England, Pamela M; Whistler, Jennifer L

    2012-08-01

    Opioid receptors, including the mu and delta opioid receptors (MOR and DOR) are important targets for the treatment of pain. Although there is mounting evidence that these receptors form heteromers, the functional role of the MOR/DOR heteromer remains unresolved. We have designed and synthesized bivalent ligands as tools to elucidate the functional role of the MOR/DOR heteromer. Our ligands (L2 and L4) are comprised of a compound with low affinity at the DOR tethered to a compound with high affinity at the MOR, with the goal of producing ligands with "tuned affinity" at MOR/DOR heteromers compared to DOR homomers. Here we show that both L2 and L4 demonstrate enhanced affinity at MOR/DOR heteromers compared to DOR homomers, thereby providing unique pharmacological tools to dissect the role of the MOR/DOR heteromer in pain.

  6. Picomolar affinity antibodies from a fully synthetic naive library selected and evolved by ribosome display.

    PubMed

    Hanes, J; Schaffitzel, C; Knappik, A; Plückthun, A

    2000-12-01

    Here we applied ribosome display to in vitro selection and evolution of single-chain antibody fragments (scFvs) from a large synthetic library (Human Combinatorial Antibody Library; HuCAL) against bovine insulin. In three independent ribosome display experiments different clusters of closely related scFvs were selected, all of which bound the antigen with high affinity and specificity. All selected scFvs had affinity-matured up to 40-fold compared to their HuCAL progenitors, by accumulating point mutations during the ribosome display cycles. The dissociation constants of the isolated scFvs were as low as 82 pM, which validates the design of the naïve library and the power of this evolutionary method. We have thus mimicked the process of antibody generation and affinity maturation with a synthetic library in a cell-free system in just a few days, obtaining molecules with higher affinities than most natural antibodies.

  7. Quantum image encryption based on generalized affine transform and logistic map

    NASA Astrophysics Data System (ADS)

    Liang, Hao-Ran; Tao, Xiang-Yang; Zhou, Nan-Run

    2016-07-01

    Quantum circuits of the generalized affine transform are devised based on the novel enhanced quantum representation of digital images. A novel quantum image encryption algorithm combining the generalized affine transform with logistic map is suggested. The gray-level information of the quantum image is encrypted by the XOR operation with a key generator controlled by the logistic map, while the position information of the quantum image is encoded by the generalized affine transform. The encryption keys include the independent control parameters used in the generalized affine transform and the logistic map. Thus, the key space is large enough to frustrate the possible brute-force attack. Numerical simulations and analyses indicate that the proposed algorithm is realizable, robust and has a better performance than its classical counterpart in terms of computational complexity.

  8. Dual-tagging system for the affinity purification of mammalian protein complexes

    SciTech Connect

    Giannone, Richard J; McDonald, W Hayes; Hurst, Gregory {Greg} B; Huang, Ying; Wu, Jun; Liu, Yie; Wang, Yisong

    2007-01-01

    Although affinity purification coupled with mass spectrometry (MS) provides a powerful tool to study protein-protein interactions, this strategy has encountered numerous difficulties when adapted to mammalian cells. Here we describe a Gateway{reg_sign}-compatible dual-tag affinity purification system that integrates regulatable expression, tetracysteine motifs, and various combinations of affinity tags to facilitate the cloning, detection, and purification of bait proteins and their interacting partners. Utilizing the human telomere binding protein TRF2 as a benchmark, we demonstrate bait protein recoveries upwards of approximately 16% from as little as 1-7 x 10{sup 7} cells and successfully identify known TRF2 interacting proteins, suggesting that our dual-tag affinity purification approach is a capable new tool for expanding the capacity to explore mammalian proteomic networks.

  9. Improving the convergence rate in affine registration of PET and SPECT brain images using histogram equalization.

    PubMed

    Salas-Gonzalez, D; Górriz, J M; Ramírez, J; Padilla, P; Illán, I A

    2013-01-01

    A procedure to improve the convergence rate for affine registration methods of medical brain images when the images differ greatly from the template is presented. The methodology is based on a histogram matching of the source images with respect to the reference brain template before proceeding with the affine registration. The preprocessed source brain images are spatially normalized to a template using a general affine model with 12 parameters. A sum of squared differences between the source images and the template is considered as objective function, and a Gauss-Newton optimization algorithm is used to find the minimum of the cost function. Using histogram equalization as a preprocessing step improves the convergence rate in the affine registration algorithm of brain images as we show in this work using SPECT and PET brain images.

  10. The Binding of Biotin to Sepharose-Avidin Column: Demonstration of the Affinity Chromatography Technique

    ERIC Educational Resources Information Center

    Landman, A. D.; Landman, N. N.

    1976-01-01

    Describes a biochemistry experiment that illustrates the methodology of affinity chromatography by attaching avidin, a glycoprotein in egg white, to a Sepharose matrix in order to bind biotin-containing proteins. (MLH)

  11. Synthesis of nonlinear discrete control systems via time-delay affine Takagi-Sugeno fuzzy models.

    PubMed

    Chang, Wen-Jer; Chang, Wei

    2005-04-01

    The affine Takagi-Sugeno (TS) fuzzy model played a more important role in nonlinear control because it can be used to approximate the nonlinear systems more than the homogeneous TS fuzzy models. Besides, it is known that the time delays exist in physical systems and the previous works did not consider the time delay effects in the analysis of affine TS fuzzy models. Hence a parallel distributed compensation based fuzzy controller design issue for discrete time-delay affine TS fuzzy models is considered in this paper. The time-delay effect is considered in the discrete affine TS fuzzy models and the stabilization issue is developed for the nonlinear time-delay systems. Finally, a numerical simulation for a time-delayed nonlinear truck-trailer system is given to show the applications of the present approach.

  12. Affinity Chromatography of Lactate Dehydrogenase: An Experiment for the Undergraduate Biochemistry Laboratory.

    ERIC Educational Resources Information Center

    Anderson, Alexander J.

    1988-01-01

    Discusses a laboratory technique of enzyme purification by affinity chromatography as part of an undergraduate biochemical methodology course. Provides preparation details of the rat muscle homogenate and reagents. Proposes column requirements and assaying information. (MVL)

  13. An Activation Force-based Affinity Measure for Analyzing Complex Networks

    PubMed Central

    Guo, Jun; Guo, Hanliang; Wang, Zhanyi

    2011-01-01

    Affinity measure is a key factor that determines the quality of the analysis of a complex network. Here, we introduce a type of statistics, activation forces, to weight the links of a complex network and thereby develop a desired affinity measure. We show that the approach is superior in facilitating the analysis through experiments on a large-scale word network and a protein-protein interaction (PPI) network consisting of ∼5,000 human proteins. The experiment on the word network verifies that the measured word affinities are highly consistent with human knowledge. Further, the experiment on the PPI network verifies the measure and presents a general method for the identification of functionally similar proteins based on PPIs. Most strikingly, we find an affinity network that compactly connects the cancer-associated proteins to each other, which may reveal novel information for cancer study; this includes likely protein interactions and key proteins in cancer-related signal transduction pathways. PMID:22355630

  14. [Reliability and validity of the Japanese revised version of the television affinity scale].

    PubMed

    Erikawa, Shigeru; Yamada, Kazunari

    2012-02-01

    The purpose of this study was to improve the Japanese version of the Television Affinity Scale (TAS), and to examine the relationship between affinity for television and viewing behavior. Data was based on a random sample of 552 people in Hachioji City (Tokyo, Japan); the response rate was 55.2%. The results revealed the following: (a) the TAS 6-item version had sufficient reliability and validity, (b) the TAS provided information which could not be explained directly by demographic factors, and (c) affinity for television was positively correlated with unplanned and non-concentrated television viewing. These results are consistent with the findings of Erikawa, Yamada, Kawabata, and Numazaki (2007). In addition, the TAS scores correlated positively with entertainment program viewing. This is consistent with the findings of Rubin (1984) that television affinity correlated with ritualized television viewing. The implications of these results for contemporary television viewing are discussed. PMID:22514907

  15. PREDICTING ER BINDING AFFINITY FOR EDC RANKING AND PRIORITIZATION: MODEL I

    EPA Science Inventory

    A Common Reactivity Pattern (COREPA) model, based on consideration of multiple energetically reasonable conformations of flexible chemicals was developed using a training set of 232 rat estrogen receptor (rER) relative binding affinity (RBA) measurements. The training set include...

  16. Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity.

    PubMed

    Boder, E T; Midelfort, K S; Wittrup, K D

    2000-09-26

    Single-chain antibody mutants have been evolved in vitro with antigen-binding equilibrium dissociation constant K(d) = 48 fM and slower dissociation kinetics (half-time > 5 days) than those for the streptavidin-biotin complex. These mutants possess the highest monovalent ligand-binding affinity yet reported for an engineered protein by over two orders of magnitude. Optimal kinetic screening of randomly mutagenized libraries of 10(5)-10(7) yeast surface-displayed antibodies enabled a >1,000-fold decrease in the rate of dissociation after four cycles of affinity mutagenesis and screening. The consensus mutations are generally nonconservative by comparison with naturally occurring mouse Fv sequences and with residues that do not contact the fluorescein antigen in the wild-type complex. The existence of these mutants demonstrates that the antibody Fv architecture is not intrinsically responsible for an antigen-binding affinity ceiling during in vivo affinity maturation.

  17. Construction of a Class of Four-Dimensional Piecewise Affine Systems with Homoclinic Orbits

    NASA Astrophysics Data System (ADS)

    Wu, Tiantian; Yang, Xiao-Song

    2016-06-01

    Based on mathematical analysis, this paper provides a methodology to ensure the existence of homoclinic orbits in a class of four-dimensional piecewise affine systems. In addition, an example is provided to illustrate the effectiveness of the method.

  18. Molecular editing of cellular responses by the high-affinity receptor for IgE.

    PubMed

    Suzuki, Ryo; Leach, Sarah; Liu, Wenhua; Ralston, Evelyn; Scheffel, Jörg; Zhang, Weiguo; Lowell, Clifford A; Rivera, Juan

    2014-02-28

    Cellular responses elicited by cell surface receptors differ according to stimulus strength. We investigated how the high-affinity receptor for immunoglobulin E (IgE) modulates the response of mast cells to a high- or low-affinity stimulus. Both high- and low-affinity stimuli elicited similar receptor phosphorylation; however, differences were observed in receptor cluster size, mobility, distribution, and the cells' effector responses. Low-affinity stimulation increased receptor association with the Src family kinase Fgr and shifted signals from the adapter LAT1 to the related adapter LAT2. LAT1-dependent calcium signals required for mast cell degranulation were dampened, but the role of LAT2 in chemokine production was enhanced, altering immune cell recruitment at the site of inflammation. These findings uncover how receptor discrimination of stimulus strength can be interpreted as distinct in vivo outcomes.

  19. Mathematical modeling of the low and high affinity arabinose transport systems in Escherichia coli.

    PubMed

    Yildirim, Necmettin

    2012-04-01

    A mathematical model was developed for the low and high affinity arabinose transport systems in E. coli. The model is a system of three ordinary differential equations and takes the dynamics of mRNAs for the araE and araFGH proteins and the internal arabinose into account. Special attention was paid to estimate the model parameters from the literature. Our analysis and simulations suggest that the high affinity transport system helps the low affinity transport system to respond to high concentration of extracellular arabinose faster, whereas the high affinity transport system responds to a small amount of extracellular arabinose. Steady state analysis of the model also predicts that there is a regime for the extracellular concentration of arabinose where the arabinose system can show bistable behavior.

  20. Purification of human copper, zinc superoxide dismutase by copper chelate affinity chromatography

    SciTech Connect

    Weslake, R.J.; Chesney, S.L.; Petkau, A.; Friesen, A.D.

    1986-05-15

    Copper, zinc superoxide dismutase was isolated from human red blood cell hemolysate by DEAE-Sepharose and copper chelate affinity chromatography. Enzyme preparations had specific activities ranging from 3400 to 3800 U/mg and recoveries were approximately 60% of the enzyme activity in the lysate. Copper chelate affinity chromatography resulted in a purification factor of about 60-fold. The homogeneity of the superoxide dismutase preparation was analyzed by sodium dodecyl sulfate-gel electrophoresis, analytical gel filtration chromatography, and isoelectric focusing.

  1. Evaluation of ligand-binding affinity using polynomial empirical scoring functions.

    PubMed

    de Azevedo, Walter Filgueira; Dias, Raquel

    2008-10-15

    Assessing protein-ligand interaction is of great importance for virtual screening initiatives in order to discover new drugs. The present work describes a set of empirical scoring functions to assess the binding affinity, involving terms for intermolecular hydrogen bonds and contact surface. The results show that our methodology works better to predict protein-ligand affinity when compared with XSCORE, a popular empirical scoring function.

  2. Exploration of dimensions of estrogen potency: parsing ligand binding and coactivator binding affinities.

    PubMed

    Jeyakumar, M; Carlson, Kathryn E; Gunther, Jillian R; Katzenellenbogen, John A

    2011-04-15

    The estrogen receptors, ERα and ERβ, are ligand-regulated transcription factors that control gene expression programs in target tissues. The molecular events underlying estrogen action involve minimally two steps, hormone binding to the ER ligand-binding domain followed by coactivator recruitment to the ER·ligand complex; this ligand·receptor·coactivator triple complex then alters gene expression. Conceptually, the potency of an estrogen in activating a cellular response should reflect the affinities that characterize both steps involved in the assembly of the active ligand·receptor·coactivator complex. Thus, to better understand the molecular basis of estrogen potency, we developed a completely in vitro system (using radiometric and time-resolved FRET assays) to quantify independently three parameters: (a) the affinity of ligand binding to ER, (b) the affinity of coactivator binding to the ER·ligand complex, and (c) the potency of ligand recruitment of coactivator. We used this system to characterize the binding and potency of 12 estrogens with both ERα and ERβ. Some ligands showed good correlations between ligand binding affinity, coactivator binding affinity, and coactivator recruitment potency with both ERs, whereas others showed correlations with only one ER subtype or displayed discordant coactivator recruitment potencies. When ligands with low receptor binding affinity but high coactivator recruitment potencies to ERβ were evaluated in cell-based assays, elevation of cellular coactivator levels significantly and selectively improved their potency. Collectively, our results indicate that some low affinity estrogens may elicit greater cellular responses in those target cells that express higher levels of specific coactivators capable of binding to their ER complexes with high affinity. PMID:21321128

  3. Assessment of Solvated Interaction Energy Function for Ranking Antibody-Antigen Binding Affinities.

    PubMed

    Sulea, Traian; Vivcharuk, Victor; Corbeil, Christopher R; Deprez, Christophe; Purisima, Enrico O

    2016-07-25

    Affinity modulation of antibodies and antibody fragments of therapeutic value is often required in order to improve their clinical efficacies. Virtual affinity maturation has the potential to quickly focus on the critical hotspot residues without the combinatorial explosion problem of conventional display and library approaches. However, this requires a binding affinity scoring function that is capable of ranking single-point mutations of a starting antibody. We focus here on assessing the solvated interaction energy (SIE) function that was originally developed for and is widely applied to scoring of protein-ligand binding affinities. To this end, we assembled a structure-function data set called Single-Point Mutant Antibody Binding (SiPMAB) comprising several antibody-antigen systems suitable for this assessment, i.e., based on high-resolution crystal structures for the parent antibodies and coupled with high-quality binding affinity measurements for sets of single-point antibody mutants in each system. Using this data set, we tested the SIE function with several mutation protocols based on the popular methods SCWRL, Rosetta, and FoldX. We found that the SIE function coupled with a protocol limited to sampling only the mutated side chain can reasonably predict relative binding affinities with a Spearman rank-order correlation coefficient of about 0.6, outperforming more aggressive sampling protocols. Importantly, this performance is maintained for each of the seven system-specific component subsets as well as for other relevant subsets including non-alanine and charge-altering mutations. The transferability and enrichment in affinity-improving mutants can be further enhanced using consensus ranking over multiple methods, including the SIE, Talaris, and FOLDEF energy functions. The knowledge gained from this study can lead to successful prospective applications of virtual affinity maturation. PMID:27367467

  4. Purification of L-( sup 3 H) Nicotine eliminates low affinity binding

    SciTech Connect

    Romm, E.; Marks, M.J.; Collins, A.C. ); Lippiello, P.M. )

    1990-01-01

    Some studies of L-({sup 3}H) nicotine binding to rodent and human brain tissue have detected two binding sites as evidenced by nonlinear Scatchard plots. Evidence presented here indicated that the low affinity binding site is not stereospecific, is not inhibited by low concentrations of cholinergic agonists and is probably due to breakdown products of nicotine since purification of the L-({sup 3}H)nicotine eliminates the low affinity site.

  5. 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.

  6. EF5 Is the High-Affinity Mg(2+) Site in ALG-2.

    PubMed

    Tanner, John J; Frey, Benjamin B; Pemberton, Travis; Henzl, Michael T

    2016-09-13

    The penta-EF-hand (PEF) protein ALG-2 (apoptosis-linked gene 2) has been implicated in several important physiological processes, including endoplasmic reticulum-Golgi vesicular transport and endosomal biogenesis/transport. ALG-2 was recently shown to harbor a metal ion-binding site with a high affinity for Mg(2+) and a low affinity for Ca(2+). We herein present the X-ray structure of Mg(2+)-bound ALG-2des23(wt). Although the C(α) trace is nearly indistinguishable from that of the Ca(2+)-free protein, the orientation of the C-terminal helix differs in the two structures. Consistent with that observation, replacement of the +x ligand in EF5, D169, with alanine eliminates high-affinity Mg(2+) binding. It also eliminates the low-affinity Ca(2+) site and lowers the affinity of the remaining Ca(2+)-binding sites, EF3 and EF1. The coordination environment in EF5 approaches ideal Mg(2+) octahedral geometry. The ligand array, consisting of three carboxylates (+x, +y, +z), a backbone carbonyl (-y), and two water molecules (-x, -z), may offer a recipe for a high-affinity, high-selectivity Mg(2+)-binding site. Sequence data for other PEF proteins indicate that select calpain large subunits, notably CAPN1 and CAPN8, may also possess a high-affinity Mg(2+)-binding site. In Mg(2+)-bound ALG-2, the carbonyl of F188 and the C-terminal carboxylate of V191 interact with the ε-ammonium group of K137 in the opposing subunit, suggesting that Mg(2+) binding could have an impact on dimerization. Interestingly, EF1 and EF3 are also occupied in the crystal, despite having modest affinity for Mg(2+). The results of a calorimetry-based analysis indicate that their Mg(2+) binding constants are 2 orders of magnitude lower than that determined for EF5. PMID:27541325

  7. Bioengineering of bacteria to assemble custom-made polyester affinity resins.

    PubMed

    Hay, Iain D; Du, Jinping; Burr, Natalie; Rehm, Bernd H A

    2015-01-01

    Proof of concept for the in vivo bacterial production of a polyester resin displaying various customizable affinity protein binding domains is provided. This was achieved by engineering various protein binding domains into a bacterial polyester-synthesizing enzyme. Affinity binding domains based on various structural folds and derived from molecular libraries were used to demonstrate the potential of this technique. Designed ankyrin repeat proteins (DARPins), engineered OB-fold domains (OBodies), and VHH domains from camelid antibodies (nanobodies) were employed. The respective resins were produced in a single bacterial fermentation step, and a simple purification protocol was developed. Purified resins were suitable for most lab-scale affinity chromatography purposes. All of the affinity domains tested produced polyester beads with specific affinity for the target protein. The binding capacity of these affinity resins ranged from 90 to 600 nmol of protein per wet gram of polyester affinity resin, enabling purification of a recombinant protein target from a complex bacterial cell lysate up to a purity level of 96% in one step. The polyester resin was efficiently produced by conventional lab-scale shake flask fermentation, resulting in bacteria accumulating up to 55% of their cellular dry weight as polyester. A further proof of concept demonstrating the practicality of this technique was obtained through the intracellular coproduction of a specific affinity resin and its target. This enables in vivo binding and purification of the coproduced "target protein." Overall, this study provides evidence for the use of molecular engineering of polyester synthases toward the microbial production of specific bioseparation resins implementing previously selected binding domains. PMID:25344238

  8. Assessment of Solvated Interaction Energy Function for Ranking Antibody-Antigen Binding Affinities.

    PubMed

    Sulea, Traian; Vivcharuk, Victor; Corbeil, Christopher R; Deprez, Christophe; Purisima, Enrico O

    2016-07-25

    Affinity modulation of antibodies and antibody fragments of therapeutic value is often required in order to improve their clinical efficacies. Virtual affinity maturation has the potential to quickly focus on the critical hotspot residues without the combinatorial explosion problem of conventional display and library approaches. However, this requires a binding affinity scoring function that is capable of ranking single-point mutations of a starting antibody. We focus here on assessing the solvated interaction energy (SIE) function that was originally developed for and is widely applied to scoring of protein-ligand binding affinities. To this end, we assembled a structure-function data set called Single-Point Mutant Antibody Binding (SiPMAB) comprising several antibody-antigen systems suitable for this assessment, i.e., based on high-resolution crystal structures for the parent antibodies and coupled with high-quality binding affinity measurements for sets of single-point antibody mutants in each system. Using this data set, we tested the SIE function with several mutation protocols based on the popular methods SCWRL, Rosetta, and FoldX. We found that the SIE function coupled with a protocol limited to sampling only the mutated side chain can reasonably predict relative binding affinities with a Spearman rank-order correlation coefficient of about 0.6, outperforming more aggressive sampling protocols. Importantly, this performance is maintained for each of the seven system-specific component subsets as well as for other relevant subsets including non-alanine and charge-altering mutations. The transferability and enrichment in affinity-improving mutants can be further enhanced using consensus ranking over multiple methods, including the SIE, Talaris, and FOLDEF energy functions. The knowledge gained from this study can lead to successful prospective applications of virtual affinity maturation.

  9. Purification of a Recombinant Polyhistidine-Tagged Glucosyltransferase Using Immobilized Metal-Affinity Chromatography (IMAC).

    PubMed

    de Costa, Fernanda; Barber, Carla J S; Pujara, Pareshkumar T; Reed, Darwin W; Covello, Patrick S

    2016-01-01

    Short peptide tags genetically fused to recombinant proteins have been widely used to facilitate detection or purification without the need to develop specific procedures. In general, an ideal affinity tag would allow the efficient purification of tagged proteins in high yield, without affecting its function. Here, we describe the purification steps to purify a recombinant polyhistidine-tagged glucosyltransferase from Centella asiatica using immobilized metal affinity chromatography. PMID:26843168

  10. 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

  11. 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.

  12. Electron Affinities, Fluoride Affinities, and Heats of Formation of the Second Row Transition Metal Hexafluorides: MF6 (M = Mo, Tc, Ru, Rh, Pd, Ag)

    SciTech Connect

    Craciun, Raluca; Long, Rebecca T.; Dixon, David A.; Christe, Karl O.

    2010-07-22

    High-level electronic structure calculations were used to evaluate reliable, self-consistent thermochemical data sets for the second row transition metal hexafluorides. The electron affinities, heats of formation, first (MF{sub 6} {yields} MF{sub 5} + F) and average M-F bond dissociation energies, and fluoride affinities of MF{sub 6} (MF{sub 6} + F{sup -} {yields} MF{sub 7}{sup -}) and MF{sub 5} (MF{sub 5} + F{sup -} {yields} MF{sub 6}{sup -}) were calculated. The electron affinities are higher than those of the corresponding third row hexafluorides, making them stronger one-electron oxidizers. The calculated electron affinities, in good agreement with the available experimental values, are 4.23 eV for MoF{sub 6}, 5.89 eV for TcF{sub 6}, 7.01 eV for RuF{sub 6}, 6.80 eV for RhF{sub 6}, 7.95 eV for PdF{sub 6}, and 8.89 eV for AgF{sub 6}. The corresponding pentafluorides are also very strong Lewis acids, although their acidities on the pF{sup -} scale are about one unit lower than those of the third row pentafluorides. The performance of a wide range of DFT exchange-correlation functionals was benchmarked by comparing them to our more accurate CCSD(T) results.

  13. GHB receptor targets in the CNS: focus on high-affinity binding sites.

    PubMed

    Bay, Tina; Eghorn, Laura F; Klein, Anders B; Wellendorph, Petrine

    2014-01-15

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects of exogenous GHB are mediated by GABA subtype B (GABAB) receptors that bind GHB with low affinity. The existence of GHB high-affinity binding sites has been known for more than three decades, but the uncovering of their molecular identity has only recently begun. This has been prompted by the generation of molecular tools to selectively study high-affinity sites. These include both genetically modified GABAB knock-out mice and engineered selective GHB ligands. Recently, certain GABA subtype A (GABAA) receptor subtypes emerged as high-affinity GHB binding sites and potential physiological mediators of GHB effects. In this research update, a description of the various reported receptors for GHB is provided, including GABAB receptors, certain GABAA receptor subtypes and other reported GHB receptors. The main focus will thus be on the high-affinity binding targets for GHB and their potential functional roles in the mammalian brain.

  14. 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

  15. Immobilizing affinity proteins to nitrocellulose: a toolbox for paper-based assay developers.

    PubMed

    Holstein, Carly A; Chevalier, Aaron; Bennett, Steven; Anderson, Caitlin E; Keniston, Karen; Olsen, Cathryn; Li, Bing; Bales, Brian; Moore, David R; Fu, Elain; Baker, David; Yager, Paul

    2016-02-01

    To enable enhanced paper-based diagnostics with improved detection capabilities, new methods are needed to immobilize affinity reagents to porous substrates, especially for capture molecules other than IgG. To this end, we have developed and characterized three novel methods for immobilizing protein-based affinity reagents to nitrocellulose membranes. We have demonstrated these methods using recombinant affinity proteins for the influenza surface protein hemagglutinin, leveraging the customizability of these recombinant "flu binders" for the design of features for immobilization. The three approaches shown are: (1) covalent attachment of thiolated affinity protein to an epoxide-functionalized nitrocellulose membrane, (2) attachment of biotinylated affinity protein through a nitrocellulose-binding streptavidin anchor protein, and (3) fusion of affinity protein to a novel nitrocellulose-binding anchor protein for direct coupling and immobilization. We also characterized the use of direct adsorption for the flu binders, as a point of comparison and motivation for these novel methods. Finally, we demonstrated that these novel methods can provide improved performance to an influenza hemagglutinin assay, compared to a traditional antibody-based capture system. Taken together, this work advances the toolkit available for the development of next-generation paper-based diagnostics. PMID:26427504

  16. ELISA-mimic screen for synthetic polymer nanoparticles with high affinity to target proteins.

    PubMed

    Yonamine, Yusuke; Hoshino, Yu; Shea, Kenneth J

    2012-09-10

    Synthetic polymer nanoparticles (NPs) that display high affinity to protein targets have significant potential for medical and biotechnological applications as protein capture agents or functional replacements of antibodies ("plastic antibodies"). In this study, we modified an immunological assay (enzyme-linked immunosorbent assay: ELISA) into a high-throughput screening method to select nanoparticles with high affinity to target proteins. Histone and fibrinogen were chosen as target proteins to demonstrate this concept. The selection process utilized a biotinylated NP library constructed with combinations of functional monomers. The screen identified NPs with distinctive functional group compositions that exhibited high affinity to either histone or fibrinogen. The variation of protein affinity with changes in the nature and amount of functional groups in the NP provided chemical insight into the principle determinants of protein-NP binding. The NP affinity was semiquantified using the ELISA-mimic assay by varying the NP concentrations. The screening results were found to correlate with solution-based assay results. This screening system utilizing a biotinylated NP is a general approach to optimize functional monomer compositions and can be used to rapidly search for synthetic polymers with high (or low) affinity for target biological macromolecules. PMID:22813352

  17. Affitins as robust tailored reagents for affinity chromatography purification of antibodies and non-immunoglobulin proteins.

    PubMed

    Béhar, Ghislaine; Renodon-Cornière, Axelle; Mouratou, Barbara; Pecorari, Frédéric

    2016-04-01

    Affinity chromatography is a convenient way of purifying proteins, as a high degree of purity can be reached in one step. The use of tags has greatly contributed to the popularity of this technique. However, the addition of tags may not be desirable or possible for the production of biopharmaceuticals. There is thus a need for tailored artificial affinity ligands. We have developed the use of archaeal extremophilic proteins as scaffolds to generate affinity proteins (Affitins). Here, we explored the potential of Affitins as ligand to design affinity columns. Affitins specific for human immunoglobulin G (hIgG), bacterial PulD protein, and chicken egg lysozyme were immobilized on an agarose matrix. The columns obtained were functional and highly selective for their cognate target, even in the presence of exogenous proteins as found in cell culture media, ascites and bacterial lysates, which result in a high degree of purity (∼95%) and recovery (∼100%) in a single step. Anti-hIgG Affitin columns withstand repetitive cycles of purification and cleaning-in-place treatments with 0.25 M NaOH as well as Protein A does. High levels of Affitin productions in Escherichia coli makes it possible to produce these affinity columns at low cost. Our results validate Affitins as a new class of tailored ligands for the affinity chromatography purification of potentially any proteins of interest including biopharmaceuticals.

  18. Integrated microfluidic approach for quantitative high-throughput measurements of transcription factor binding affinities

    PubMed Central

    Glick, Yair; Orenstein, Yaron; Chen, Dana; Avrahami, Dorit; Zor, Tsaffrir; Shamir, Ron; Gerber, Doron

    2016-01-01

    Protein binding to DNA is a fundamental process in gene regulation. Methodologies such as ChIP-Seq and mapping of DNase I hypersensitive sites provide global information on this regulation in vivo. In vitro methodologies provide valuable complementary information on protein–DNA specificities. However, current methods still do not measure absolute binding affinities. There is a real need for large-scale quantitative protein–DNA affinity measurements. We developed QPID, a microfluidic application for measuring protein–DNA affinities. A single run is equivalent to 4096 gel-shift experiments. Using QPID, we characterized the different affinities of ATF1, c-Jun, c-Fos and AP-1 to the CRE consensus motif and CRE half-site in two different genomic sequences on a single device. We discovered that binding of ATF1, but not of AP-1, to the CRE half-site is highly affected by its genomic context. This effect was highly correlated with ATF1 ChIP-seq and PBM experiments. Next, we characterized the affinities of ATF1 and ATF3 to 128 genomic CRE and CRE half-site sequences. Our affinity measurements explained that in vivo binding differences between ATF1 and ATF3 to CRE and CRE half-sites are partially mediated by differences in the minor groove width. We believe that QPID would become a central tool for quantitative characterization of biophysical aspects affecting protein–DNA binding. PMID:26635393

  19. Food and value motivation: Linking consumer affinities to different types of food products.

    PubMed

    de Boer, Joop; Schösler, Hanna

    2016-08-01

    This study uses the consumer affinity concept to examine the multiple motives that may shape consumers' relationships with food. The concept was applied in a study on four broad product types in the Netherlands, which cover a wide range of the market and may each appeal to consumers with different affinities towards foods. These product types may be denoted as 'conventional', 'efficient', 'gourmet' and 'pure'. A comparative analysis, based on Higgins' Regulatory Focus Theory, was performed to examine whether food-related value motivations could explain different consumer affinities for these product types. The affinities of consumers were measured by means of a non-verbal, visual presentation of four samples of food products in a nationwide survey (n = 742) among consumers who were all involved in food purchasing and/or cooking. The affinities found could be predicted fairly well from a number of self-descriptions relating to food and eating, which expressed different combinations of type of value motivation and involvement with food. The analysis demonstrated the contrasting role of high and low involvement as well as the potential complementarity of promotion- and prevention-focused value motivation. It is suggested that knowledge of the relationships between product types, consumer affinities and value motivation can help improve the effectiveness of interventions that seek to promote healthy and sustainable diets in developed countries. PMID:27046434

  20. Immobilizing affinity proteins to nitrocellulose: a toolbox for paper-based assay developers.

    PubMed

    Holstein, Carly A; Chevalier, Aaron; Bennett, Steven; Anderson, Caitlin E; Keniston, Karen; Olsen, Cathryn; Li, Bing; Bales, Brian; Moore, David R; Fu, Elain; Baker, David; Yager, Paul

    2016-02-01

    To enable enhanced paper-based diagnostics with improved detection capabilities, new methods are needed to immobilize affinity reagents to porous substrates, especially for capture molecules other than IgG. To this end, we have developed and characterized three novel methods for immobilizing protein-based affinity reagents to nitrocellulose membranes. We have demonstrated these methods using recombinant affinity proteins for the influenza surface protein hemagglutinin, leveraging the customizability of these recombinant "flu binders" for the design of features for immobilization. The three approaches shown are: (1) covalent attachment of thiolated affinity protein to an epoxide-functionalized nitrocellulose membrane, (2) attachment of biotinylated affinity protein through a nitrocellulose-binding streptavidin anchor protein, and (3) fusion of affinity protein to a novel nitrocellulose-binding anchor protein for direct coupling and immobilization. We also characterized the use of direct adsorption for the flu binders, as a point of comparison and motivation for these novel methods. Finally, we demonstrated that these novel methods can provide improved performance to an influenza hemagglutinin assay, compared to a traditional antibody-based capture system. Taken together, this work advances the toolkit available for the development of next-generation paper-based diagnostics.