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.     

NMR study of the complexation of boric acid with catechol (1,2-dihydroxybenzene)

The boric acid–catechol system in aqueous solutions has been studied by means of ¹¹B, ¹H and ¹³C NMR over wide ranges of concentration and pH. ¹¹B spectra show a striking difference between the complexation possibilities of dilute and more concentrated solutions of boric acid. Both the 1:1 and 1:2 complexes can be simultaneously observed by ¹³C NMR.     

Development of an integrated optic oxygen sensor using a novel, generic platform

This paper describes the development of a generic platform for enhanced, integrated optic sensors based on fluorescence detection. The platform employs a novel optical configuration in order to achieve enhanced performance and has inherent multianalyte detection capability. The sensor element comprises a multimode ridge waveguide that has been patterned with an analyte-sensitive fluorescent spot, which is excited directly using a LED. The platform was applied to the detection of gaseous oxygen as a proof of principle. The sol-gel-derived sensor spots were doped with an oxygen-sensitive fluorescent dichlororuthenium dye complex and intensity-based calibration data were generated from the oxygen-dependent waveguide output. The sensor achieved a LOD of 0.62% and a resolution of less than 0.96% gaseous oxygen, which compares favourably with a similar, recently reported system. This device highlights the combination of inexpensive rapid prototyping techniques and a dedicated sensor enhancement strategy that together facilitate the production of an effective prototype sensor platform.     

RCM of tripeptide dienes containing a chiral vinylcyclopropane moiety: impact of different Ru-based catalysts on the stereochemical integrity of the macrocyclic products

[structures: see text] Tripeptide dienes containing an (1R,2S)-vinyl aminocyclopropylcarboxylate residue were cyclized to beta-strand scaffolds under ring-closing metathesis (RCM). Conformational factors, ligand effects, and reaction conditions were evaluated. A protocol was developed for the efficient synthesis of 15-membered ring peptides in high diastereomeric purity. These peptides are key synthetic precursors to antiviral agents that target the hepatitis C virus and represent the first class of clinically validated pharmaceutical agents that are synthesized in large scale using RCM.     

Thermally stimulated current (TSC) study of the Tg and Tll transitions in anionic polystyrenes

The thermally stimulated current (TSC) technique has been used to investigate three anionic polystyrenes of M?_n 17,000, 71,700, and 1.55 × 10⁶, i.e., M < M_c, M > M_c, and M ? M_c, where M_c is the entanglement molecular weight. A current maximum near T_g designated T_Mg, has relaxation times which follow an Arrhenius equation. A second current maximum at T > T_g appears to be the T_ll process and is designated T_Mll. Relaxation times for it follow a Vogel equation. T_Mg and T_Mll vary with molecular weight, increasing below M_c and leveling off above M_c at a temperature of about 170°C. Values of T_Mg and T_Mll are compared with values of T_g and T_ll obtained from torsional braid analysis, which involves melt flow; and with differential-scanning-calorimetric values on fused films, where there is no transport of polymer. It is concluded from such cross-comparisons that TSC, at least for polystyrene, is a quasistatic test which may involve microscopic viscosity. Macroscopic viscosity does not play a role. The ratio T_Mll/T_Mg is in the range 1.10–1.16, similar to T_ll/T_g values by other methods. A few comments about T_ll in atactic poly(methyl methacrylate) by the TSC method are given.     

Induction of Wavelength-Dependent Photochemistry in Bilirubins by Serum Albumin

Summary.  Quantum yields for Z⇄E photoisomerization of the natural bi-chromophore bilirubin IXα in ammoniacal methanol solution were found to vary with excitation wavelength, whereas no variation (within experimental error) was observed for xanthobilirubic acid and for symmetrically substituted bilirubins (bilirubin IIIα, bilirubin XIIIα, and mesobilirubin XIIIα) in the same solvent. The quantum yield for Z⇄E photoisomerization of xanthobilirubic acid bound to human serum albumin was also invariant with excitation wavelength. In contrast, quantum yields for Z⇄E photoisomerization of the symmetrically-substituted bilirubins did show marked excitation wavelength dependency when they were bound to human serum albumin. These results show that quantum yields for Z⇄E photoisomerization of bilirubins are markedly sensitive to protein binding and to the nature of pyrrole ring β-substituents. They also demonstrate that wavelength-dependent photochemistry is characteristic of bilirubins with non-identical pyrromethenone chromophores, as expected from exciton coupling theory, and that complexation with albumin induces wavelength-dependent photochemistry in symmetrically substituted bilirubins. Received January 13, 1998. Accepted January 27, 1998     

Optical Properties of Micro-patterned Silver Nanoparticle Substrates

In this paper, we describe a novel technique for depositing metal nanoparticles (NPs) on a planar substrate whereby the NPs are micro-patterned on the surface by a simple stamp-printing procedure. The method exploits the attractive force between negatively charged colloidal metal NPs and positively-charged polyelectrolyte layers which have been selectively deposited on the surface. Using this technique, large uniform areas of patterned metal NPs, with different plasmonic properties, were achieved by optimisation of the stamping process. We report the observation of unusual fluorescence emission from these structures. The emission was measured using epifluorescence microscopy. Fluorescence lifetime behaviour was also measured. Furthermore, the μ-patterned NPs exhibited blinking behaviour under 469 nm excitation and the fluorescence spectrum was multi-peaked. It has been established that the fluorescence is independent of the plasmon resonance properties of the NPs. As well as optimising the novel NP μ-patterning technique, this work discusses the origin and characteristics of the anomalous fluorescence behaviour in order to characterise and minimise this unwanted background contribution in the use of metal NPs for plasmonic enhancement of fluorescence for optical biochip applications.