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The multicopper enzyme nitrous oxide reductase (N2OR) catalyzes the final step of denitrification, the two-electron reduction of N2O to N-2. This enzyme is a functional homodimer containing two different multicopper sites: CuA and CuZ. CuA is a binuclear copper site that transfers electrons to the tetranuclear copper sulfide CuZ, the catalytic site. In this study, Pseudomonas nautica cytochrome C-552 was identified as the physiological electron donor. The kinetic data show differences when physiological and artificial electron donors are compared [cytochrome vs methylviologen (MV)]. In the presence of cytochrome c(552), the reaction rate is dependent on the ET reaction and independent of the N2O concentration. With MV, electron donation is faster than substrate reduction. From the study of cytochrome c(552) concentration dependence, we estimate the following kinetic parameters: K-mc512 = 50.2 +/- 9.0 mu M and V-maxc551 1.8 +/- 10.6 units/mg. The N2O concentration dependence indicates a K-mN2O of 14.0 +/- 2.9 mu M using MV as the electron donor. The pH effect on the kinetic parameters is different when MV or cytochrome c(552) is used as the electron donor (pK(a) = 6.6 or 8.3, respectively). The kinetic study also revealed the hydrophobic nature of the interaction, and direct electron transfer studies showed that CuA is the center that receives electrons from the physiological electron donor. The formation of the electron transfer complex was observed by H-1 NMR protein-protein titrations and was modeled with a molecular docking program (BiGGER). The proposed docked complexes corroborated the ET studies giving a large number of solutions in which cytochrome c(552) is placed near a hydrophobic patch located around the CuA center.

1st ISA Forum report

In this paper will be discussed different types of scenarios and the aims for using scenarios. Normaly they are being used by organisations due to the need to anticipate processes, to support policy-making and to understand the complexities of relations. Such organisations can be private companies, R&D organisations and networks of organisations, or even by some public administration institutions. Some cases will be discussed as the methods for ongoing scenario-building process (Shell Internacional). Scenarios should anticipate possible relations among social actors as in the Triple Helix Model, and is possible to develop strategic intelligence in the innovation process that would enable the construction of scenarios. Such processes can be assessed. The focus will be made in relation to the steps chosen for the WORKS scenarios. In this case is there a model of work changes that can be used for foresight? Differences according to sectors were found, as well on other dimensions. Problems of assessment are analysed with specific application to the scenario construction methods.

No abstract is available for this item.

WORKS final conference report

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Membrane-bound nitrate reductase from Marinobacter hydrocarbonoclasticus 617 can be solubilized in either of two ways that will ultimately determine the presence or absence of the small (I) subunit. The enzyme complex (NarGHI) is composed of three subunits with molecular masses of 130, 65, and 20 kDa. This enzyme contains approximately 14 Fe, 0.8 Mo, and 1.3 molybdopterin guanine dinucleotides per enzyme molecule. Curiously, one heme b and 0.4 heme c per enzyme molecule have been detected. These hemes were potentiometrically characterized by optical spectroscopy at pH 7.6 and two noninteracting species were identified with respective midpoint potentials at E(m) = + 197 mV (heme c) and-4.5 mV (heme b). Variable-temperature (4-120 K) X-band electron paramagnetic resonance (EPR) studies performed on both as-isolated and dithionite-reduced nitrate reductase showed, respectively, an EPR signal characteristic of a {[}3Fe-4S](+) cluster and overlapping signals associated with at least three types of {[}4Fe-4S](+) centers. EPR of the as-isolated enzyme shows two distinct pH-dependent Mo(V) signals with hyperfine coupling to a solvent-exchangeable proton. These signals, called ``lowpH'' and ``high-pH,'' changed to a pH-independent Mo(V) signal upon nitrate or nitrite addition. Nitrate addition to dithionite-reduced samples at pH 6 and 7.6 yields some of the EPR signals described above and a new rhombic signal that has no hyperfine structure. The relationship between the distinct EPR-active Mo(V) species and their plausible structures is discussed on the basis of the structural information available to date for closely related membrane-bound nitrate reductases.

Direct electrochemical response was first time observed for the redox centers of Desulfovibrio gigas [NiFe]-Hase, in non-turnover conditions, by cyclic voltammetry, in solution at glassy carbon electrode. The activation of the enzyme was achieved by reduction with H(2) and by electrochemical control and electrocatalytic activity was observed. The inactivation of the [NiFe]-Hase was also attained through potential control. All electrochemical data was obtained in the absence of enzyme inhibitors. The results are discussed in the context of the proposed mechanism currently accepted for activation/inactivation of [NiFe]-Hases. (C) 2008 Elsevier B.V. All rights reserved.

Metalloenzymes control enzymatic activity by changing the characteristics of the metal centers where catalysis takes place. The conversion between inactive and active states can be tuned by altering the coordination number of the metal site, and in some cases by an associated conformational change. These processes will be illustrated using heme proteins (cytochrome c nitrite reductase, cytochrome c peroxidase and cytochrome cd(1) nitrite reductase), non-heme proteins (superoxide reductase and [ NiFe]-hydrogenase), and copper proteins (nitrite and nitrous oxide reductases) as examples. These examples catalyze electron transfer reactions that include atom transfer, abstraction and insertion.

Adenylate kinase (AK) mediates the reversible transfer of phosphate groups between the adenylate nucleotides and contributes to the maintenance of their constant cellular level, necessary for energy metabolism and nucleic acid synthesis. The AK were purified from crude extracts of two sulfate-reducing bacteria (SRB), Desulfovibrio (D.) gigas NCIB 9332 and Desulfovibrio desulfuricans ATCC 27774, and biochemically and spectroscopically characterised in the native and fully cobalt- or zinc-substituted forms. These are the first reported adenylate kinases that bind either zinc or cobalt and are related to the subgroup of metal-containing AK found, in most cases, in Gram-positive bacteria. The electronic absorption spectrum is consistent with tetrahedral coordinated cobalt, predominantly via sulfur ligands, and is supported by EPR. The involvement of three cysteines in cobalt or zinc coordination was confirmed by chemical methods. Extended X-ray absorption fine structure (EXAFS) indicate that cobalt or zinc are bound by three cysteine residues and one histidine in the metal-binding site of the ``LID{''} domain. The sequence (129)Cys-X(5)-His-X(15)-Cys-X(2)-Cys of the AK from D. gigas is involved in metal coordination and represents a new type of binding motif that differs from other known zinc-binding sites of AK. Cobalt and zinc play a structural role in stabilizing the LID domain. (C) 2008 Elsevier Inc. All rights reserved.

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This work reports for the first time a resonance Raman study of the mixed-valence and fully reduced forms of Paracoccus pantotrophus bacterial cytochrome c peroxidase. The spectra of the active mixed-valence enzyme show changes in the structure of the ferric peroxidatic heme compared to the fully oxidized enzyme; these differences are observed upon reduction of the electron-transferring heme and upon full occupancy of the calcium site. For the mixed-valence form in the absence of Ca2+, the peroxidatic heme is six-coordinate and low-spin on the basis of the frequencies of the structure-sensitive Raman lines: the enzyme is inactive. With added Ca2+, the peroxidatic heme is five-coordinate high-spin and active. The calcium-dependent spectral differences indicate little change in the conformation of the ferrous electron-transferring heme, but substantial changes in the conformation of the ferric peroxidatic heme. Structural changes associated with Ca2+ binding are indicated by spectral differences in the structure-sensitive marker lines, the out-of-plane low-frequency macrocyclic modes, and the vibrations associated with the heme substituents of that heme. The Ca2+-dependent appearance of a strong gamma(15) saddling-symmetry mode for the mixed-valence form is consistent with a strong saddling deformation in the active peroxidatic heme, a feature seen in the Raman spectra of other peroxidases. For the fully reduced form in the presence of Ca2+, the resonance Raman spectra show that the peroxidatic heme remains high-spin.

Portugal had only very few foresight exercises on the automobile sector, and the most recent one was a survey held in a project on work organisation systems in the automobile industry, its recent historical paths and the special strategies of location of companies (the WorTiS project). This involved several teams with different disciplinary backgrounds and from two Portuguese universities. The provisional main results of the first round of a Delphi survey held in Portugal on the automotive sector were already published, but a further analysis was not yet done. This foresight survey was done under the WorTiS project, developed in 2004 by IET – Research Centre on Enterprise and Work Innovation (at FCT-UNL), and financed by the Portuguese Ministry of Science and Technology. Some of this experience on foresight analysis is also been transferred to other projects, namely the WORKS project on work organisation restructuring in the knowledge society that received the support from EC and still is running. The majority of experts considered having an average of less knowledge in almost all the scenario topics presented. This means that information on the automotive industry is not spread enough among academics or experts in related fields (regional scientists, innovation economists, engineers, sociologists). Some have a good knowledge but in very specialised fields. Others have expertise on foresight, or macroeconomics, or management sciences, but feel insecure on issues related with futures of automobile sector. Nevertheless, we considered specially the topics where the experts considered themselves to have some knowledge. There were no “irrelevant” topics considered as such by the expert panel. There are also no topics that are not considered a need for co-operation. The lack of technological infrastructures was not considered as a hindered factor for the accomplishment of any scenario. The experts’ panel considered no other international competence besides US, Jap

Portugal had only very few foresight exercises on the automobile sector, and the most recent one was a survey held in a project on work organisation systems in the automobile industry, its recent historical paths and the special strategies of location of companies (the WorTiS project). This involved several teams with different disciplinary backgrounds and from two Portuguese universities. The provisional main results of the first round of a Delphi survey held in Portugal on the automotive sector were already published, but a further analysis was not yet done. This foresight survey was done under the WorTiS project, developed in 2004 by IET – Research Centre on Enterprise and Work Innovation (at FCT-UNL), and financed by the Portuguese Ministry of Science and Technology. Some of this experience on foresight analysis is also been transferred to other projects, namely the WORKS project on work organisation restructuring in the knowledge society that received the support from EC and still is running. The majority of experts considered having an average of less knowledge in almost all the scenario topics presented. This means that information on the automotive industry is not spread enough among academics or experts in related fields (regional scientists, innovation economists, engineers, sociologists). Some have a good knowledge but in very specialised fields. Others have expertise on foresight, or macroeconomics, or management sciences, but feel insecure on issues related with futures of automobile sector. Nevertheless, we considered specially the topics where the experts considered themselves to have some knowledge. There were no “irrelevant” topics considered as such by the expert panel. There are also no topics that are not considered a need for co-operation. The lack of technological infrastructures was not considered as a hindered factor for the accomplishment of any scenario. The experts' panel considered no other international competence besides US, Japan

Nesta dissertação apresenta-se um motor em disco polifásico inovador bem como uma estratégia de controlo com base no método de variação de velocidade por comutação do número de pares de pólos. A configuração das bobinas aliada à escolha das correntes e tensões que se injectam nas bobinas dos estatores, permite comutar electronicamente o número de pólos do motor entre 2, 4 6 e 8 pólos, conseguindo-se controlar a característica binário?velocidade do motor. O motor em disco possui a bobinagem feita em cobre com dois semi-estatores, em que quando utiliza o rotor em alumínio (com condutividade diferente de zero) comporta-se como um motor de indução convencional. Quando se substitui o rotor em alumínio por um constituído por um supercondutor de alta temperatura (SAT), o dispositivo comporta-se como um motor de histerese. O princípio de funcionamento do motor em disco convencional é baseado na indução de força electromotrizes no rotor e, consequentemente, uma vez que o alumínio é bom condutor eléctrico, correntes eléctricas induzidas, originadas por haver um campo magnético variável que é criado pelos semi-estatores. O comportamento deste tipo de motores, no que diz respeito a principais características (como o binário?velocidade para os diferentes números de pares de pólos), circuito equivalente de Steinmetz, entre outras teorias associadas é já conhecido há bastante tempo. O princípio de funcionamento do motor SAT é diferente do apresentado anteriormente, funciona com base na dinâmica de vórtices e devido ao facto de aparecer o fenómeno de ancoragem de fluxo (flux pinning) nos supercondutores de alta temperatura. Como o campo magnético varia, então o disco roda. Este motor tem um princípio de funcionamento muito mais complexo que o motor de indução sendo a obtenção do modelo do motor SAT complicada. A obtenção do modelo do motor SAT não é abordado nesta dissertação. Os comportamentos e modos de operação do motor com disco de alumínio e em materiais SAT são simulados através de um programa comercial de elementos finitos, nesta dissertação, sendo a supercondutividade simulada com base na relação entre o campo eléctrico e a densidade de corrente pela lei da potenciação (E-J power law). Com as simulações pretende-se comparar o rendimento electromecânico de ambos os motores.

Nesta dissertação apresenta-se um motor em disco polifásico inovador bem como uma estratégia de controlo com base no método de variação de velocidade por comutação do número de pares de pólos. A configuração das bobinas aliada à escolha das correntes e tensões que se injectam nas bobinas dos estatores, permite comutar electronicamente o número de pólos do motor entre 2, 4 6 e 8 pólos, conseguindo-se controlar a característica binário?velocidade do motor. O motor em disco possui a bobinagem feita em cobre com dois semi-estatores, em que quando utiliza o rotor em alumínio (com condutividade diferente de zero) comporta-se como um motor de indução convencional. Quando se substitui o rotor em alumínio por um constituído por um supercondutor de alta temperatura (SAT), o dispositivo comporta-se como um motor de histerese. O princípio de funcionamento do motor em disco convencional é baseado na indução de força electromotrizes no rotor e, consequentemente, uma vez que o alumínio é bom condutor eléctrico, correntes eléctricas induzidas, originadas por haver um campo magnético variável que é criado pelos semi-estatores. O comportamento deste tipo de motores, no que diz respeito a principais características (como o binário?velocidade para os diferentes números de pares de pólos), circuito equivalente de Steinmetz, entre outras teorias associadas é já conhecido há bastante tempo. O princípio de funcionamento do motor SAT é diferente do apresentado anteriormente, funciona com base na dinâmica de vórtices e devido ao facto de aparecer o fenómeno de ancoragem de fluxo (flux pinning) nos supercondutores de alta temperatura. Como o campo magnético varia, então o disco roda. Este motor tem um princípio de funcionamento muito mais complexo que o motor de indução sendo a obtenção do modelo do motor SAT complicada. A obtenção do modelo do motor SAT não é abordado nesta dissertação. Os comportamentos e modos de operação do motor com disco de alumínio e em materiais SAT são simulados através de um programa comercial de elementos finitos, nesta dissertação, sendo a supercondutividade simulada com base na relação entre o campo eléctrico e a densidade de corrente pela lei da potenciação (E-J power law). Com as simulações pretende-se comparar o rendimento electromecânico de ambos os motores.

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