Superoxide reductase (SOR) is a metalloprotein containing a non-heme iron centre, responsible for the scavenging of superoxide radicals in the cell. The crystal structure of Treponema pallidum (Tp) SOR was determined using soft X-rays and synchrotron radiation. Crystals of the oxidized form were obtained using poly(ethylene glycol) and MgCl2 and diffracted beyond 1.55 A resolution. The overall architecture is very similar to that of other known SORs but TpSOR contains an N-terminal domain in which the desulforedoxin-type Fe centre, found in other SORs, is absent. This domain conserves the beta-barrel topology with an overall arrangement very similar to that of other SOR proteins where the centre is present. The absence of the iron ion and its ligands, however, causes a decrease in the cohesion of the domain and some disorder is observed, particularly in the region where the metal would be harboured. The C-terminal domain exhibits the characteristic immunoglobulin-like fold and harbours the Fe(His)4(Cys) active site. The five ligands of the iron centre are well conserved despite some disorder observed for one of the four molecules in the asymmetric unit. The participation of a glutamate as the sixth ligand of some of the iron centres in Pyrococcus furiosus SOR was not observed in TpSOR. A possible explanation is that either X-ray photoreduction occurred or there was a mixture of redox states at the start of data collection. In agreement with earlier proposals, details in the TpSOR structure also suggest that Lys49 might be involved in attraction of superoxide to the active site.

Desulfovibrio vulgaris Hildenborough cytochrome c(3) contains four hemes in a low-spin state with bis-histidinyl coordination. High-spin forms of cytochrome c(3) can be generated by protonation of the axial ligands in order to probe spin equilibrium (low-spin/high-spin). The spin alterations occurring at acid pH, the associated changes in redox potentials, as well as the reactivity towards external ligands were followed by the conjunction of square wave voltammetry and UV-visible, CD, NMR and EPR spectroscopies. These processes may be used for modelling the action of enzymes that use spin equilibrium to promote enzyme activity and reactivity towards small molecules.

A new family of compounds able to promote host-guest interactions with specific molecules (e.g., cyanuric and parabanic acids) and to coordinate metal ions, namely Zn(II) and Cu(II), has been synthesized and fully characterized. The new probes derive from the attachment of two methylaminopyrene units to the carbonyl precursor 2,6-bis(2-formylphenoxymethyl)pyridine. Its signalling properties result from the fluorescence emission properties, which reveal the existence of intramolecular excimer formation. The compounds have showed to be highly sensitive to the solvent and hydrogen ion concentration of the medium. Depending on these, different monomer-to-excimer fluorescence ratio is displayed by the two probes. The compound with a single pyrene unit revealed absence of excimer formation and was used as model compound. The overall results are discussed on the basis of the studied probes as potentially revealing molecular movements, off-on-off fluorescent photoinduced electron transfer (PET), host-guest interactions with specific compounds and of sensing metal ions. (c) 2005 Elsevier B.V. All rights reserved.

Among the Krebs cycle components, just citrate enhances the fluorescence of a new bi(brachial) lariat aza-crown containing appended naphthalene fluorophores.

No abstract included.

n/a

n/a

Herein the preparation of molecularly imprinted polymers (MIPs) using supercritical fluid technology is evaluated. Poly(diethylene glycol dimethacrylate), polyDEGDMA, was synthesised in supercritical carbon dioxide (scCO2) using a carboxylic acid end-capped perfluoropolyether oil as stabiliser. Polymerisations were carried out in the presence of different concentrations of two different template drug molecules, salicylic acid and acetylsalicylic acid. Results suggest that molecular imprinted polymers were successfully prepared by supercritical polymerisation and then impregnated with the template in order to prepare controlled release systems.

This report begins with some general information and analysis of policy and regulation that were the subjects of discussion and exchange in the policy pillar in the first phase of WORKS. The second section is a synthesis of country information on general principles and trends of policy and policy enforcement. This is followed by a summary of sector information for the sectors chosen by the qualitative pillar to be the objects of empirical analysis. The last summarises research questions and dimensions to be guidelines for carrying out case studies and capturing the relevance and effects of policy and institutions at the workplace.

n/a

Superoxide reductase is a 14 kDa metalloprotein containing a catalytic non-haem iron centre [Fe(His)4Cys]. It is involved in defence mechanisms against oxygen toxicity, scavenging superoxide radicals from the cell. The oxidized form of Treponema pallidum superoxide reductase was crystallized in the presence of polyethylene glycol and magnesium chloride. Two crystal forms were obtained depending on the oxidizing agents used after purification: crystals grown in the presence of K3Fe(CN)6 belonged to space group P2(1) (unit-cell parameters a = 60.3, b = 59.9, c = 64.8 A, beta = 106.9 degrees) and diffracted beyond 1.60 A resolution, while crystals grown in the presence of Na2IrCl6 belonged to space group C2 (a = 119.4, b = 60.1, c = 65.6 A, beta = 104.9 degrees) and diffracted beyond 1.55 A. A highly redundant X-ray diffraction data set from the C2 crystal form collected on a copper rotating-anode generator (lambda = 1.542 A) clearly defined the positions of the four Fe atoms present in the asymmetric unit by SAD methods. A MAD experiment at the iron absorption edge confirmed the positions of the previously determined iron sites and provided better phases for model building and refinement. Molecular replacement using the P2(1) data set was successful using a preliminary trace as a search model. A similar arrangement of the four protein molecules could be observed.

A nitrate reductase was solubilized with Triton X-100 from the membranes of Pseudomonas chlororaphis DSM 50135 grown microaerobically in the presence of nitrate. Like other membrane-bound nitrate reductases, it contains three subunits, of 129, 66 (64) and 24 kDa, referred to in the literature as alpha, beta and gamma, respectively. Electrocatalytic studies revealed that only the membrane-bound, not the solubilized form of the enzyme, can accept electrons from a menaquinone analog, menadione, whereas both forms can accept electrons from methylviologen. The isolated enzyme possesses several iron-sulfur clusters and a molybdopterin guanine dinucleotide active center. The iron-sulfur clusters can be grouped in two classes according to their redox properties, the high-potential and low-potential clusters. In the as-isolated enzyme, two forms of the molybdenum center, high- and low-pH, are detectable by electron paramagnetic resonance spectroscopy. The low-pH form shows a hyperfine splitting due to a proton, suggesting the presence of an -OHx ligand. Dithionite reduces the Mo(V) center to Mo(W) and subsequent reoxidization with nitrate originates a new Mo(V) signal, identical to the oxidized low-pH form but lacking its characteristic hyperfine splitting. The isolated preparation also contains heme c (in a sub-stoichiometric amount) with the ability to relay electrons to the molybdenum center, suggesting that this nitrate reductase may contain heme c instead of the heme b usually found in this class of enzymes. (c) 2005 Elsevier B.V. All rights reserved.