Monitoring protein interactions in the living cell through the fluorescence decays of the cyan fluorescent protein.

Using fluorescence lifetime microspectroscopy and imaging techniques, we have studied the fluorescence of cyan fluorescent protein (CFP) transiently expressed in HEK-293 cells, in the presence or absence of its fluorescence resonance energy transfer (FRET) partner, yellow fluorescent protein (YFP). When the two proteins are attached through a 27-amino-acid linker, a 33 % average efficiency of intramolecular energy transfer is accurately determined inside the cell. Additionally, we observe a systematic quenching of the CFP fluorescence with increasing levels of protein expression. This quenching cannot be accounted for by formation of the previously described dimer of GFP-related proteins, since its magnitude is unchanged when the fluorescent proteins carry the mutation A206K shown to dissociate this dimer in vitro. Even when the intracellular protein concentration largely exceeds the in vitro dissociation constant of the dimer, self-association remains undetectable, either between free proteins or intramolecularly within the CFP-YFP construct. Instead, the detailed concentration effects are satisfactorily accounted for by a model of intermolecular, concentration-dependent energy transfer, arising from molecular proximity and crowding. In the case of CFP alone, we suggest that self-quenching could result from a pseudo-homo FRET mechanism between different, spectrally shifted emissive forms of the protein. These phenomena require careful consideration in intracellular FRET studies.     

Synthesis of P1 aspartate-based peptide acyloxymethyl and fluoromethyl ketones as inhibitors of interleukin-1β-converting enzyme

Improved procedures have been developed for the synthesis of P1 aspartate-based 2,6-dichlorobenzoyloxymethyl ketone 1 and fluoromethyl ketone 2, the prodrugs of two potent ICE-inhibitors. 1 was prepared from (R)-trans-4,5-O-isopropylidene-4,5-dihydroxy-2-pentenecarboxylic acid ethyl ester; 2 was obtained via a nitro-aldol condensation as key step from in situ generated fluoroacetaldehyde.     

Electron entanglement near a superconductor and bell inequalities

Near the interface between a normal metal and a superconductor, Cooper pairs penetrate into the normal side, giving rise to the proximity effect. The two electrons of these pairs have entangled spin and orbital degrees of freedom. Nonlocal features of quantum mechanics can be probed by separating these two electrons. This is achieved with a fork geometry with two normal leads containing either spin- or energy-selective filters. A signature of entanglement can be detected by measuring the positive noise cross-correlations in this fork. In the case of energy filters, Bell-inequality checks constitute a definite probe of entanglement. We formulate Bell-type inequalities in terms of current-current cross-correlations associated with contacts with varying magnetization orientations. We find maximal violation (as in photons) when a superconductor is the particle source.     

Size-selected synthesis of PtRu nano-catalysts: reaction and size control mechanism

A rapid synthesis method for the preparation of PtRu colloids and their subsequent deposition on high surface area carbons is presented. The reaction mechanism is shown to involve the oxidation of the solvent, ethylene glycol, to mainly glycolic acid or, depending on the pH, its anion, glycolate, while the Pt(+IV) and Ru(+III) precursor salts are reduced. Glycolate acts as a stabilizer for the PtRu colloids and the glycolate concentration, and hence the size of the resulting noble metal colloids is controlled via the pH of the synthesis solution. Carbon-supported PtRu catalysts of controlled size can be prepared within the range of 0.7-4 nm. Slow scan X-ray diffraction and high-resolution transmission electron microscopy show the PtRu catalysts to be crystalline. The Ru is partly dissolved in the face-centered cubic Pt lattice, but the catalysts also consist of a separate, hexagonal Ru phase. The PtRu catalysts appear to be of the same composition independent of the catalyst size in the range of 1.2-4 nm. Particular PtRu catalysts prepared in this work display enhanced activities for the CH(3)OH electro-oxidation reaction when compared to two commercial catalysts.     

Polyferrocenes: metallopolymers with tunable and high refractive indices

The refractive index, molar refraction and Abbe number of polyferrocene derivatives are reported and the values indicate that these materials are very promising for a range of photonics applications.     

Dynamic optimization and event location in atmospheric chemistry

A model coupling differential equations and an optimization problem with equality and inequality constraints is described. The first order optimality conditions are coupled with the differential equations to form a differential-algebraic system. Inequality constraints for the optimization variables induce discontinuities in the first derivatives of the solution in time. Tracking techniques based on sensitivity analysis and second order multistep methods are proposed to locate the discontinuities. Application to the dynamics of organic atmospheric aerosol particles is given to illustrate the detection of the activation of constraints. (© 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)     

A numerical study of vorticity layers in a two-dimensional stratified shear flow

A numerical study is conducted to find out the conditions of occurrence of a secondary Kelvin-Helmholtz instability in the thin layers (referred to as baroclinic layers) that form in a stably-stratified shear layer. For this purpose, three high resolution calculations of a moderately stratified shear layer have been carried out, at a fixed Reynolds number. The wavelength of the initial perturbation is progressively increased, starting from the fundamental wavelength predicted by linear stability theory up to twice this fundamental wavelength. The baroclinic layer of the flow is shown to lengthen and destabilize progressively from one calculation to the other, eventually bearing a secondary Kelvin-Helmholtz instability. The structure and dynamics of the baroclinic layers of the three calculations are examined in the frame of a theoretical model proposed by Corcos and Sherman ([1]). An excellent agreement with the predictions of this model have been found. We next show that the stability of the layer is controlled by the large-scale Kelvin-Helmholtz vortex, via the strain field that it induces in the stagnation point region of the layer. A consequence of this study is that secondary Kelvin-Helmholtz instabilities are fostered by the pairing of primary Kelvin-Helmholtz vortices in a strongly-stratified shear layer.

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Sommario E stato condotto uno studio numerico per trovare le condizioni in cui insorge una instabilità secondaria di Kelvin-Helmholtz negli strati sottili che si formano in uno strato di scorrimento stabilmente stratificato. A questo scopo sono state effettuate tre simulazioni ad alta risoluzione a fissato numero di Reynolds e stratificazione bassa. La lunghezza d'onda della perturbazione iniziale è stata progressivamente aumentata dalla lunghezza fondamentale predetta dalla teoria lineare della stabilità fino a due volte questa stessa lunghezza. È stato osservato che da una simulazione all'altra lo strato baroclino del flusso si allunga e si destabilizza progressivamente, generando eventualmente un'instabilità di Kelvin-Helmholtz secondaria. Utilizzando il modello teorico proposto da Corcos e Sherman (1976), per le tre simulazioni sono state analizzate la struttura e la dinamica dello strato baroclino. È stato trovato un accordo eccellente con le predizioni di questo modello. È stato in seguito mostrato che la stabilità dello strato è controllato dai vortici di Kelvin-Helmholtz di larga scala attraverso il campo di deformazione che inducono nella regione del punto di ristagno dello strato. Una conseguenza di questo studio è che le instabilità secondarie di Kelvin-Helmholtz sono forzate dall'accoppiamento dei vortici primari in uno strato di scorrimento fortemente stratificato.

     

Cationic Biphotonic Lanthanide Luminescent Bioprobes Based on Functionalized Cross-Bridged Cyclam Macrocycles

Cationic lanthanide complexes are generally able to spontaneously internalize into living cells. Following our previous works based on a diMe-cyclen framework, a second generation of cationic water-soluble lanthanide complexes based on a constrained cross-bridged cyclam macrocycle functionalized with donor-π-conjugated picolinate antennas was prepared with europium(III) and ytterbium(III). Their spectroscopic properties were thoroughly investigated in various solvents and rationalized with the help of DFT calculations. A significant improvement was observed in the case of the Eu³⁺ complex, while the Yb³⁺ analogue conserved photophysical properties in aqueous solvent. Two-photon (2P) microscopy imaging experiments on living T24 human cancer cells confirmed the spontaneous internalization of the probes and images with good signal-to-noise ratio were obtained in the classic NIR-to-visible configuration with the Eu³⁺ luminescent bioprobe and in the NIR-to-NIR with the Yb³⁺ one.     

The Role of the Nucleotides in the Insertion of the bis-Molybdopterin Guanine Dinucleotide Cofactor into apo-Molybdoenzymes

The role of the GMP nucleotides of the bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor of the DMSO reductase family has long been a subject of discussion. The recent characterization of the bis-molybdopterin (bis-Mo-MPT) cofactor present in the E. coli YdhV protein, which differs from bis-MGD solely by the absence of the nucleotides, now enables studying the role of the nucleotides of bis-MGD and bis-MPT cofactors in Moco insertion and the activity of molybdoenzymes in direct comparison. Using the well-known E. coli TMAO reductase TorA as a model enzyme for cofactor insertion, we were able to show that the GMP nucleotides of bis-MGD are crucial for the insertion of the bis-MGD cofactor into apo-TorA.     

Reevaluating the Solution Photophysics of Tetraphenylethylene at the Origin of their Aggregation-Induced Emission Properties

Although tetraphenylethylene (TPE) and its derivatives have been the most commonly used building blocks in the construction of molecules with aggregation-induced emission (AIE) properties, no absolute consensus exists regarding the mechanisms at the origin of the phenomenon. Restriction of intramolecular rotations (RIR) of peripheral phenyls has historically been a dominant paradigm, which has served as a valuable guideline in the molecular engineering of AIEgens. Yet, an increasing number of recent works have established that photoisomerization or photocyclization may actively participate in the nonradiative dissipation of the excitation energy. In this paper, the first experimental evaluation of the quantum efficiencies of these different processes is reported, and photoisomerization is shown to be by far the dominant photophysical pathway in solution, accounting for virtually all nonradiative decay of the molecule‘s excited state in degassed solution.