Respiratory nitric oxide reductase (NOR) was purified from membrane extract of Pseudomonas (Ps.) nautica cells to homogeneity as judged by polyacrylamide gel electrophoresis. The purified protein is a heterodimer with subunits of molecular masses of 54 and 18 kDa. The gene encoding both subunits was cloned and sequenced. The amino acid sequence shows strong homology with enzymes of the cNOR class. Iron/heme determinations show that one heme c is present in the small subunit (NORC) and that approximately two heme b and one non-heme iron are associated with the large subunit (NORB), in agreement with the available data for enzymes of the cNOR class. Mossbauer characterization of the as-purified, ascorbate-reduced, and dithionite-reduced enzyme confirms the presence of three heme groups (the catalytic heme b(3) and the electron transfer heme b and heme c) and one redox-active non-heme Fe (Fe(B)). Consistent with results obtained for other cNORs, heme c and heme b in Ps. nautica cNOR were found to be low-spin while Fe(B) was found to be high-spin. Unexpectedly, as opposed to the presumed high-spin state for heme b(3), the Mossbauer data demonstrate unambiguously that heme b(3) is, in fact, low-spin in both ferric and ferrous states, suggesting that heme b(3) is six-coordinated regardless of its oxidation state. EPR spectroscopic measurements of the as-purified enzyme show resonances at the g approximately 6 and g approximately 2-3 regions very similar to those reported previously for other cNORs. The signals at g = 3.60, 2.99, 2.26, and 1.43 are attributed to the two charge-transfer low-spin ferric heme c and heme b. Previously, resonances at the g approximately 6 region were assigned to a small quantity of uncoupled high-spin Fe(III) heme b(3). This assignment is now questionable because heme b(3) is low-spin. On the basis of our spectroscopic data, we argue that the g = 6.34 signal is likely arising from a spin-spin coupled binuclear center comprising the low-spin Fe(III) heme b(3) and the high-spin Fe(B)(III). Activity assays performed under various reducing conditions indicate that heme b(3) has to be reduced for the enzyme to be active. But, from an energetic point of view, the formation of a ferrous heme-NO as an initial reaction intermediate for NO reduction is disfavored because heme [FeNO](7) is a stable product. We suspect that the presence of a sixth ligand in the Fe(II)-heme b(3) may weaken its affinity for NO and thus promotes, in the first catalytic step, binding of NO at the Fe(B)(II) site. The function of heme b(3) would then be to orient the Fe(B)-bound NO molecules for the formation of the N-N bond and to provide reducing equivalents for NO reduction.

Saturation transfer difference (STD) NMR has emerged as one of the most popular ligand-based NMR techniques for the study of protein−ligand interactions. The success of this technique is a consequence of its robustness and the fact that it is focused on the signals of the ligand, without any need of processing NMR information about the receptor and only using small quantities of nonlabeled macromolecule. Moreover, the attractiveness of this experiment is also extendable to the classroom. In the context of a practical NMR class, this experiment is ideal to illustrate some fundamental NMR concepts, such as the nuclear Overhauser effect and relaxation in a multidisciplinary context, bridging chemistry and biochemistry with a taste of medicinal chemistry.

We use the readily available human serum albumin (HSA), 6-d,l-methyl-tryptophan (6-CH3-Trp), and 7- d,l-methyl-tryptophan (7-CH3-Trp) to introduce the STD-NMR experiment and to illustrate its applicability for ligand screening, mapping of binding moieties, and determination of the dissociation constant, in a context that can be explored or adapted to the student’s course level and topic (chemistry or biochemistry). We also cover the most important theoretical aspects of the STD experiment, calling attention to some of its limitations and drawbacks without a complex theoretical approach.

A forelimb of a new sauropod dinosaur (Angolatitan adamastor n. gen. et sp.) from the Late Turonian of Iembe (Bengo Province) represents the first dinosaur discovery in Angola, and is one of the few occurrences of sauropod dinosaurs in sub-Saharan Africa collected with good chronological controls. The marginal marine sediments yielding the specimen are reported to be late Turonian in age and, thus it represents a non-titanosaurian sauropod in sub-Saharan Africa at a time taken to be dominated by titanosaurian forms. Moreover, Angolatitan adamastor is the only basal Somphospondyli known in the Late Cretaceous which implies in the existence of relict forms in Africa.

Although the Late Jurassic of Portugal has provided abundant dinosaur fossils, material from the Early Cretaceous is scarce. This paper reports new cranial and postcranial material of the theropod dinosaur Baryonyx walkeri found in the Barremian (Papo Seco Formation) of Portugal. This specimen, found at Praia das Aguncheiras, Cabo Espichel, consists of a partial dentary, isolated teeth, pedal ungual, two calcanea, presacral and caudal vertebrae, fragmentary pubis, scapula, and rib fragments. It represents the most complete spinosaurid yet discovered in the Iberian Peninsula and the most complete dinosaur from the Early Cretaceous of Portugal. This specimen is confidently identified as a member of Baryonychinae due to the presence of conical teeth with flutes and denticles in a dentary rosette. The specimen ML1190 shares the following characteristics with Baryonyx walkeri: enamel surface with small (nearly vertical) wrinkles, variable denticle size along the carinae, 6–7 denticles per mm, wrinkles forming a 45 degree angle near the carinae, and tooth root longer than crown. In addition, dubious taxa based on teeth morphology such as Suchosaurus cultridens (Owen, 1840–1845), and Suchosaurus girardi (Sauvage 1897–98; Antunes & Mateus 2003) are discussed, based on comparisons with well-known material such as Baryonyx walkeri Charig & Milner, 1986. Suchosaurus cultridens and S. girardi are considered as nomina dubia due to the lack of diagnostic apomorphies, but both specimens are referred to Baryonychinae incertae sedis.

The effect of different mild post-annealing treatments in air, at 270 degrees C, for 4-6 min, on the optical, electrical, structural and chemical properties of copper sulphide (CuxS) thin films deposited at room temperature are investigated. CuxS films, 70nm thick, are deposited on glass substrates by vacuum thermal evaporation from a Cu2S:S (50:50 wt.%) sulphur rich powder mixture. The as-deposited highly conductive crystalline CuS (covellite) films show high carrier concentration (similar to 10(22) cm(-3)), low electrical resistivity (similar to 10(-4) Omega cm) and inconclusive p-type conduction. After the mild post-annealing, these films display increasing values of resistivity (similar to 10(-3) to similar to 10(-2) Omega cm) with annealing time and exhibit conclusive p-type conduction. An increase of copper content in CuxS phases towards the semiconductive Cu2S (chalcocite) compound with annealing time is reported, due to re-evaporation of sulphur from the films. However, the latter stoichiometry was not obtained, which indicates the presence of vacancies in the Cu lattice. In the most resistive films a Cu2O phase is also observed, diminishing the amount of available copper to combine with sulphur, and therefore the highest values of optical transmittance are reached (65%). The appearance on the surface of amorphous sulphates with annealing time increase is also detected as a consequence of sulphur oxidation and replacement of sulphur with oxygen. All annealed films are copper deficient in regards to the stoichiometric Cu2S and exhibit stable p-type conductivity. (C) 2011 Elsevier B.V. All rights reserved.

The effect of different mild post-annealing treatments in air, at 270 degrees C, for 4-6 min, on the optical, electrical, structural and chemical properties of copper sulphide (CuxS) thin films deposited at room temperature are investigated. CuxS films, 70nm thick, are deposited on glass substrates by vacuum thermal evaporation from a Cu2S:S (50:50 wt.%) sulphur rich powder mixture. The as-deposited highly conductive crystalline CuS (covellite) films show high carrier concentration (similar to 10(22) cm(-3)), low electrical resistivity (similar to 10(-4) Omega cm) and inconclusive p-type conduction. After the mild post-annealing, these films display increasing values of resistivity (similar to 10(-3) to similar to 10(-2) Omega cm) with annealing time and exhibit conclusive p-type conduction. An increase of copper content in CuxS phases towards the semiconductive Cu2S (chalcocite) compound with annealing time is reported, due to re-evaporation of sulphur from the films. However, the latter stoichiometry was not obtained, which indicates the presence of vacancies in the Cu lattice. In the most resistive films a Cu2O phase is also observed, diminishing the amount of available copper to combine with sulphur, and therefore the highest values of optical transmittance are reached (65%). The appearance on the surface of amorphous sulphates with annealing time increase is also detected as a consequence of sulphur oxidation and replacement of sulphur with oxygen. All annealed films are copper deficient in regards to the stoichiometric Cu2S and exhibit stable p-type conductivity. (C) 2011 Elsevier B.V. All rights reserved.

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