Understanding ligand–protein interactions in affinity membrane chromatography for antibody purification

Affinity chromatography with Protein A beads has become the conventional unit operation for the primary capture of monoclonal antibodies. However, Protein A activated supports are expensive and ligand leakage is an issue to be considered. In addition, the limited production capabilities of the chromatographic process drive the research towards feasible alternatives. The use of synthetic ligands as Protein A substitutes has been considered in this work. Synthetic ligands, that mimic the interaction between Protein A and the constant fragment (Fc) of immunoglobulins, have been immobilized on cellulosic membrane supports. The resulting affinity membranes have been experimentally characterized with pure immunoglobulin G (IgG). The effects of the membrane support and of the spacer arm on the ligand–ligate interaction have been studied in detail. Experimental data have been compared with molecular dynamic simulations with the aim of better understanding the interaction mechanisms. Molecular dynamic simulations were performed in explicit water, modelling the membrane as a matrix of overlapped glucopyranose units. Electrostatic charges of the ligand and spacer were calculated through ab initio methods to complete the force field used to model the membrane. The simulations enabled to elucidate how the interactions of surface, spacer and ligand with IgG, contribute to the formation of the bond between protein and affinity membrane.     

Optimization of enzymatic production of biodiesel from castor oil in organic solvent medium

We studied the production of fatty acid ethyl esters from castor oil using n-hexane as solvent and two commercial lipases, Novozym 435 and Lipozyme IM, as catalysts. For this purpose, a Taguchi experimental design was adopted considering the following variables: temperature (35–65°C), water (0–10 wt/wt%), and enzyme (5–20 wt/wt%) concentrations and oil-to-ethanol molar ratio (1∶3 to 1∶10). An empirical model was then built so as to assess the main and cross-variable effects on the reaction conversion and also to maximize biodiesel production for each enzyme. For the system containing Novozym 435 as tatalyst the maximum conversion obtained was 81.4% at 65°C, enzyme concentration of 20 wt/wt%, water concentration of 0 wt/wt%, and oil-to-ethanol molar ratio of 1∶10. When the catalyst was Lipozyme IM, a conversion as high as 98% was obtained at 65°C, enzyme concentration of 20 wt/wt%, water concentration of 0 wt/wt%, and oil-to-ethanol molar ratio of 1∶3.     

Rearrangement of N-Aryl-2-Vinylaziridines to Benzoazepines and Dihydropyrroles: A Synthetic and Theoretical Study

Herein we report the one-pot synthesis of several N-heterocyclic compounds by rearrangement reactions of N-aryl-2-vinylaziridines. The optimization of the synthetic methodology employed allowed us to obtain differently substituted 2,5-dihydro-1H-benzo[b]azepines in good yields and purities. The relationship between the nature of the starting N-aryl-2-vinylaziridine and the obtained N-heterocycle was also investigated. Finally, to rationalize all the experimental results reported in this paper a theoretical study was performed that casts light on the reaction mechanism.     

Relation between Different Types of Global Attractors of Set-Valued Nonautonomous Dynamical Systems

The article is devoted to the study of the relation between forward and pullback attractors of set-valued nonautonomous dynamical systems (cocycles). Here it is proved that every compact global forward attractor is also a pullback attractor of the set-valued nonautonomous dynamical system. The inverse statement, generally speaking, is not true, but we prove that every global pullback attractor of an α-condensing set-valued cocycle is always a local forward attractor. The obtained general results are applied while studying periodic and homogeneous systems. We give also a new criterion of the absolute asymptotic stability of nonstationary discrete linear inclusions. Dedicated to our friend Professor Enrico Primo Tomasini on the occasion of his 55th birthdayMathematics Subject Classifications (2000) Primary: 34C35, 34D20, 34D40, 34D45, 58F10,58F12, 58F39; secondary: 35B35, 35B40.     

Optical characterization of the phosphate glasses containing pair transition ions

The paper deals with optical and electronic properties of the aluminophosphate glasses containing Fe–Mn and Fe–Cr ion pairs in different concentration. The influence of the mixed alkali ions over the electronic properties has been investigated. The optical behavior (optical transmission) of the glass samples has been studied by UV-VIS spectroscopy and the refractive index dependency on wavelength has been discussed. The transmission spectra show features specific for the doping transition ions (TM), revealing different oxidation states of iron (Fe²⁺/Fe³⁺), manganese (Mn²⁺/Mn³⁺) and chromium (Cr³⁺/Cr⁶⁺) in the vitreous network. Mössbauer spectroscopy offers information regarding the TM oxidation states, redox processes and the iron coordination symmetry in the vitreous network. In the case of Fe–Mn doped glasses, the percentage of Fe²⁺ is about 40% and a doubled iron content leads to an increasing of Fe²⁺ percentage up to 53%. The replacing of lithium ions by natrium ions (mixed alkali effect) provides an increasing of the Fe²⁺ percentage up to 56%. The occurrence of the tetrahedral or octahedral symmetry of Fe²⁺ ions bonded by O^2? ions depends on the transition ion nature and Li⁺/Na⁺ ratio. Infrared absorption spectra of the pair transition ions-doped aluminophosphate glasses reveal optical phonons specific for the phosphate glass matrix.     

Adsorption of water on reconstructed rutile TiO2(011)-(2 x 1): Ti=O double bonds and surface reactivity

Recent combined experimental and theoretical studies (Beck et al., Phys. Rev. Lett. 2004, 93, 036104) have provided evidence for Ti=O double-bonded titanyl groups on the reconstructed rutile TiO(2)(011)-(2 x 1) surface. The adsorption of water on the same surface is now investigated to further probe the properties of these groups, as well as to confirm their existence. Ultraviolet photoemission experiments show that water is adsorbed in molecular form at a sample temperature of 110 K. At the same time, the presence of a 3sigma state in the photoemission spectra and work function measurements indicate a significant amount of hydroxyls within the first monolayer of water. At room temperature, scanning tunneling microscopy (STM) suggests that dissociated water is present, and about 30% of the surface active sites are hydroxylated. These findings are well explained by total energy density functional theory calculations and Car-Parrinello molecular dynamics simulations for water adsorption on the titanyl model of TiO(2)(011)-(2 x 1). The theoretical results show that a mixed molecular/dissociative layer is the most stable configuration in the monolayer regime at low temperatures, while complete dissociation takes place at 250 K. The arrangement of the protonated mono-coordinated oxygens in the mixed molecular/dissociated layer is consistent with the observed short-range order of the hydroxyls in the STM images.     

Backward and forward-looking expectations in a chaotic cobweb model

In this study we research the dynamics shown by a cobweb-type model with linear demand, non-invertible supply function and with forward-looking expectations associated to backward looking ones. The study of the dynamics exhibited by our model will show how the presence of forward-looking expectations represents a stabilising factor: As the weight attributed to the above mentioned foreseeing form increases, the system becomes less and less complex until it only generates orbits converging either to the fixed point or to a cycle-2. It is particularly interesting to analyse at the same time the role played by the two forms of expectations considered, both of which contribute to market stabilization.     

Enantioselective Aza‐Michael Addition of Imides by Using an Integrated Strategy Involving the Synthesis of a Family of Multifunctional Catalysts,Usage of Multiple Catalysis,and Rational Design of Experiment

A challenging asymmetric reaction (aza‐Michael addition of imides to enones) has been optimized through an integrated approach involving the synthesis of a family of organocatalysts, multiple catalysis (usage of additives), and finally with rational exploration of the chemical space by the application of the experiment design.     

A phosphotyrosine-imprinted polymer receptor for the recognition of tyrosine phosphorylated peptides

Hyperphosphorylation at tyrosine is commonly observed in tumor proteomes and, hence, specific phosphoproteins or phosphopeptides could serve as markers useful for cancer diagnostics and therapeutics. The analysis of such targets is, however, a challenging task, because of their commonly low abundance and the lack of robust and effective preconcentration techniques. As a robust alternative to the commonly used immunoaffinity techniques that rely on phosphotyrosine(pTyr)-specific antibodies, we have developed an epitope-imprinting strategy that leads to a synthetic pTyr-selective imprinted polymer receptor. The binding site incorporates two monourea ligands placed by preorganization around a pTyr dianion template. The tight binding site displayed good binding affinities for the pTyr template, in the range of that observed for corresponding antibodies, and a clear preference for pTyr over phosphoserine (pSer). In further analogy to the antibodies, the imprinted polymer was capable of capturing short tyrosine phosphorylated peptides in the presence of an excess of their non-phosphorylated counterparts or peptides phosphorylated at serine.