Oxidation of Zn(Cys)4 zinc finger peptides by O2 and H2O2: products, mechanism and kinetics

The reactivity of a series of Zn(Cys)(4) zinc finger model peptides towards H(2)O(2) and O(2) has been investigated. The oxidation products were identified by HPLC and ESI-MS analysis. At pH<7.5, the zinc complexes and the free peptides are oxidised to bis-disulfide-containing peptides. Above pH 7.5, the oxidation of the zinc complexes by H(2)O(2) also yields sulfinate- and sulfonate-containing overoxidised peptides. At pH 7.0, monitoring of the reactions between the zinc complexes and H(2)O(2) by HPLC revealed the sequential formation of two disulfides. Several techniques for the determination of the rate constant for the first oxidation step corresponding to the attack of H(2)O(2) by the Zn(Cys)(4) site have been compared. This rate constant can be reliably determined by monitoring the oxidation by HPLC, fluorescence, circular dichroism or absorption spectroscopy in the presence of excess ethyleneglycol bis(2-aminoethyl ether)tetraacetic acid. In contrast, monitoring of the release of zinc with 4-(2-pyridylazo)resorcinol or of the thiol content with 5,5'-dithiobis(2-nitrobenzoate) did not yield reliable values of this rate constant for the case in which the formation of the second disulfide is slower than the formation of the first. The kinetic measurements clearly evidence a protective effect of zinc on the oxidation of the cysteines by both H(2)O(2) and O(2), which points to the fact that zinc binding diminishes the nucleophilicity of the thiolates. In addition, the reaction between the zinc finger and H(2)O(2) is too slow to consider zinc fingers as potential sensors for H(2)O(2) in cells.     

Biosensors elaborated on gold nanoparticles, a PM-IRRAS characterisation of the IgG binding efficiency

This work is focused on studying the grafting of gold nanoparticles (Np) on a cystamine self-assembled monolayer on gold, in order to build sensitive immunosensors. The synthesis and deposition of gold nanoparticles, 13 and 55 nm sizes, were characterised by combining Polarisation Modulation Infrared Reflection-Absorption Spectroscopy (PM-IRRAS), X-ray Photoelectron Spectroscopy (XPS) Surface Enhanced Raman Scattering (SERS), and Atomic Force Microscopy (AFM) which all indicated the formation of a dispersed layer of nanoparticles. This observation is explained by the compromise between the high reactivity of amine-terminated layers towards gold, and interparticle repulsions. Nps were then functionalised with antibody probes, and the recognition by an anti-rIgG was assayed both on planar and Np gold surfaces.The important result is that nanoparticles of 55 nm are preferable for the following reasons: they enable to build a denser and well dispersed layer and they increase both the number of receptors (IgGs) and their accessibility. Beside these geometric improvements, a net enhancement of the Raman signal was observed on the 55 nm nanoparticle layer, making this new platform promising for optical detection based biosensors.     

[H3tren] templated iron fluorides; synthesis, crystal structures and Mössbauer studies

The hydrothermal synthesis, using tris-(2-ethylamino)amine (tren) as a template, and the crystal structures of three new hybrid iron fluorides, (H₃O)₂·[H₃tren]₂·(FeF₆)₂·(FeF₅(H₂O))·2H₂O (I), [H₃tren]₂·(FeF₆)₂·(FeF₂(H₂O)₄)·8H₂O (II) and [H₃tren]₂·(FeF₆)·(F)₃·H₂O (III), are reported. I, II, and III are triclinic (P-1), monoclinic (P2₁/c) and orthorhombic (I222), respectively. The structure of I is built up from isolated FeF₆ and FeF₅(H₂O) distorted octahedra separated by triprotonated [H₃tren]³⁺ cations, disordered H₃O⁺ cations and H₂O molecules. In II, Fe^IIIF₆ and neutral [Fe^IIF₂(H₂O)₄] octahedra form, together with [H₃tren]³⁺ cations, infinite (100) layers separated by extra water molecules. The structure of III consists of isolated and disordered FeF₆ octahedra, fluoride anions F⁻ connected to [H₃tren]³⁺ cations and extra fluoride anions F⁻ disordered with H₂O molecules. All [H₃tren]³⁺ cations have a “spider” type conformation. ⁵⁷Fe Mössbauer characterization shows that +III valence state can only be considered for iron cations in I and III and preliminary Mössbauer results are consistent with the presence of both +II and +III valences for iron cations in II, in agreement with the crystallographic results.     

Sorption thermodynamics of CO2, CH4, and their mixtures in the ITQ-1 zeolite as revealed by molecular simulations

The thermodynamic properties and siting of carbon dioxide and methane sorbed in the siliceous form of zeolite MCM-22, ITQ-1, were studied by means of grand canonical Monte Carlo simulation. ITQ-1 comprises two independent pore systems of different geometry. It was found to be CO(2)-selective toward CO(2)/CH(4) gas mixtures, its equilibrium selectivity being distinctly higher in its sinusoidal channel pore system than in the large cavity system over a wide range of pressures starting from the Henry law regime, at the three temperatures considered. A maximum in selectivity is observed at low temperature, high pressure, and methane-rich gas-phase composition.     

A multianalyte ELISA for immunochemical screening of sulfonamide,fluoroquinolone and ß-lactam antibiotics in milk samples using class-selective bioreceptors

A multianalyte ELISA has been developed for the simultaneous determination of the most frequently used antibiotic families in the veterinary field following the typical planar microarray configuration, where the identity of the target analyte is encoded by its location in the detection platform (Master et al. in Drug Discovery Today 11:1007-1011, 2006). To accomplish this aim, two individual enzyme-linked immunosorbent assays for sulfonamide and fluoroquinolone antibiotics and an enzyme-linked receptor assay for ss-lactam antibiotics have been combined. The strategy uses microplates coated with the corresponding haptenized proteins in specific sections of the microplate. The samples are mixed with a cocktail containing the bioreagents, and distributed in the wells of the microplate. Identification of the antibiotic present in a particular sample is consequently accomplished by detecting a positive response on the corresponding microplate section. Since the bioreceptors used show a wide recognition of the congeners of each antibiotic family, the multianalyte method is able to detect more than 25 different antibiotics from the three most important antibiotic families. The detectability reached in full-fat milk samples is below the European maximum residue limits. The accuracy and reliability of this multiplexed bioanalytical method have been demonstrated by analyzing blind spiked samples.     

Un modèle hyperbolique diphasique bi-fluide en milieu poreux

We introduce an hyperbolic two-fluid two-pressure model to compute unsteady two-phase flows in porous media. The closure laws comply with the entropy inequality, and a unique set of jump conditions holds within each field. To cite this article: J.-M. Hérard, C. R. Mecanique 336 (2008).     

Noncovalent associations in fluorous fluids

Perfluorocarbons (PFCs) are emerging as a new type of liquid phase in which molecular recognition processes can effectively take place. The combination of perfluorocarbons (PFCs) and noncovalent associations, mostly hydrogen bonds, ion pairing, halogen bonds or coordination bonds, has already been successfully exploited for applications in organic synthesis (catalyst recycling, by-product removal), electrochemical sensing, selective extraction/titration processes or to prepare gels. Due to the extreme solvophobic effect in PFCs, the least polar existing fluids, noncovalent associations tend to be enhanced. For instance, quantitative data on the increase in association strength occurring in PFCs have recently been reported for ion-pairing interactions or encapsulation processes. Moreover, several examples show that confining a receptor in a fluorous phase leads to recognition processes with improved selectivity.     

Glycerol and glycerol carbonate as ultraviscous solvents for mixture analysis by NMR

NMR of weakly polar analytes in an apolar ultraviscous solvent has recently been proposed for mixture analysis as a pertinent alternative to the DOSY experiment. The present article reports the first use of glycerol and glycerol carbonate as polar solvents for the NMR analysis of a model mixture of dipeptides. This work demonstrates the high potentiality of these solvents for the analysis of mixtures made of polar and potentially bioactive compounds. Medium-sized molecules slowly reorient in glycerol and glycerol carbonate under particular temperature conditions, so that solute resonances may show spin diffusion in NOESY spectra, thus opening the way to mixture analysis. Glycerol and glycerol carbonate have turned out to be ultraviscous solvents of choice for the individualization of four structurally close mixed dipeptides: Leu-Val, Leu-Tyr, Gly-Tyr and Ala-Tyr by means of 1D and 2D NOESY experiments. Selective sample excitation and signal detection were implemented to eliminate the intense proton signals of the non-deuterated solvents. Moreover, the recording of a multiplet selective 2D NOESY-TOCSY has shown that the analytical power of NMR in highly viscous solvents is not limited to the extraction of mixture component 1D subspectra but may also yield some supplementary information about atom connectivity within components.