Export 2155 results:
Sort by: Author Title Type [ Year  (Desc)]
1996
Gu, ZJ, Dong J, Allan CB, Choudhury SB, Franco R, Moura JJG, Legall J, Przybyla AE, Roseboom W, Albracht SPJ, Axley MJ, Scott RA, Maroney MJ.  1996.  Structure of the Ni sites in hydrogenases by X-ray absorption spectroscopy. Species variation and the effects of redox poise, Nov 13. Journal of the American Chemical Society. 118:11155-11165., Number 45 AbstractWebsite

Structural information obtained from the analysis of nickel K-edge X-ray absorption spectroscopic data of [NiFe]hydrogenases from Desulfovibrio gigas, Thiocapsa roseopersicina, Desulfovibrio desulfuricans (ATCC 27774), Escherichia coli (hydrogenase-1), Chromatium vinosum, and Alcaligenes eutrophus H16 (NAD(+)-reducing, soluble hydrogenase), poised in different redox states, is reported. The data allow the active-site structures of enzymes from several species to be compared, and allow the effects of redox poise on the structure of the nickel sites to be examined. In addition, the structure of the nickel site obtained from recent crystallographic studies of the D. gigas enzyme (Volbeda, A.; Charon, M.-H.; Piras, C.; Hatchikian, E. C.; Frey, M.; Fontecilla-Camps, J. C. Nature 1995, 373, 580-587) is compared with the structural features obtained from the analysis of XAS data from the same enzyme. The nickel sites of all but the oxidized (as isolated) sample of A. eutrophus hydrogenase are quite similar. The nickel K-edge energies shift 0.9-1.5 eV to lower energy upon reduction from oxidized (forms A and B) to fully reduced forms. This value is comparable with no more than a one-electron metal-centered oxidation state change. With the exception of T. roseopersicina hydrogenase, most of the edge energy shift (-0.8 eV) occurs upon reduction of the oxidized enzymes to the EPR-silent intermediate redox level (SI). Analysis of the XANES features assigned to 1s-->3d electronic transitions indicates that the shift in energy that occurs for reduction of the enzymes to the SI level may be attributed at least in part to an increase in the coordination number from five to six. The smallest edge energy shift is observed for the T. roseopersicina enzyme, where the XANES data indicate that the nickel center is always six-coordinate. With the exception of the oxidized sample of A. eutrophus hydrogenase, the EXAFS data are dominated by scattering from S-donor ligands at similar to 2.2 Angstrom. The enzyme obtained from T. roseopersicina also shows evidence for the presence of O,N-donor ligands. The data from A. eutrophus hydrogenase are unique in that they indicate that a significant structural change occurs upon reduction of the enzyme. EXAFS data obtained from the oxidized (as isolated) A. eutrophus enzyme indicate that the EXAFS is dominated by scattering from 3-4 N,O-donor atoms at 2.06(2) Angstrom, with contributions from 2-3 S-donor ligands at 2.35(2) Angstrom. This changes upon reduction to a more typical nickel site composed of similar to 4 S-donor ligands at a Ni-S distance of 2.19(2) Angstrom. Evidence for the presence of atoms in the 2.4-2.9 Angstrom distance range is found in most samples, particularly the reduced enzymes (SI, form C, and R). The analysis of these data is complicated by the fact that it is difficult to distinguish between S and Fe scattering atoms at this distance, and by the potential presence of both S and another metal atom at similar distances. The results of EXAFS analysis are shown to be in general agreement with the published crystal structure of the D. gigas enzyme.

Coelho, AV, Matias PM, Sieker LC, Morais J, Carrondo MA, Lampreia J, Costa C, Moura JJ, Moura I, Legall J.  1996.  Preliminary crystallographic analysis and further characterization of a dodecaheme cytochrome c from Desulfovibrio desulfuricans ATCC 27774, Nov 1. Acta Crystallogr D Biol Crystallogr. 52:1202-8., Number Pt 6 AbstractWebsite

Dodecaheme cytochrome c has been purified from Desulfovibrio (D.) desulfuricans ATCC 27774 cells grown under both nitrate and sulfate-respiring conditions. Therefore, it is likely to play a role in the electron-transfer system of both respiratory chains. Its molecular mass (37768 kDa) was determined by electrospray mass spectrometry. Its first 39 amino acids were sequenced and a motif was found between amino acids 32 and 37 that seems to exist in all the cytochromes of the c(3) type from sulfate-reducing bacteria sequenced at present. The midpoint redox potentials of this cytochrome were estimated to be -68, -120, -248 and -310 mV. Electron paramagnetic resonance spectroscopy of the oxidized cytochrome shows several low-spin components with a g(max) spreading from 3.254 to 2.983. Two crystalline forms were obtained by vapour diffusion from a solution containing 2% PEG 6000 and 0.25-0.75 M acetate buffer pH = 5.5. Both crystals belong to monoclinic space groups: one is P2(1), with a = 61.00, b = 106.19, c = 82.05 A, beta = 103.61 degrees, and the other is C2 with a = 152.17, b = 98.45, c = 89.24 A, beta = 119.18 degrees. Density measurements of the P2(1) crystals suggest that there are two independent molecules in the asymmetric unit. Self-rotation function calculations indicate, in both crystal forms, the presence of a non-crystallographic axis perpendicular to the crystallographic twofold axis. This result and the calculated values for the volume per unit molecular weight of the C2 crystals suggest the presence of two or four molecules in the asymmetric unit.

Devreese, B, Tavares P, Lampreia J, Van Damme N, Legall J, Moura JJ, Van Beeumen J, Moura I.  1996.  Primary structure of desulfoferrodoxin from Desulfovibrio desulfuricans ATCC 27774, a new class of non-heme iron proteins, May 6. FEBS Lett. 385:138-42., Number 3 AbstractWebsite

The primary structure of desulfoferrodoxin from Desulfovibrio desulfuricans ATCC 27774, a redox protein with two mononuclear iron sites, was determined by automatic Edman degradation and mass spectrometry of the composing peptides. It contains 125 amino acid residues of which five are cysteines. The first four, Cys-9, Cys-12, Cys-28 and Cys-29, are responsible for the binding of Center I which has a distorted tetrahedral sulfur coordination similar to that found in desulforedoxin from D. gigas. The remaining Cys-115 is proposed to be involved in the coordination of Center II, which is probably octahedrally coordinated with predominantly nitrogen/oxygen containing ligands as previously suggested by Mossbauer and Raman spectroscopy.

McGinnity, DF, Devreese B, Prazeres S, Van Beeumen J, Moura I, Moura JJ, Pettigrew GW.  1996.  A single histidine is required for activity of cytochrome c peroxidase from Paracoccus denitrificans, May 10. J Biol Chem. 271:11126-33., Number 19 AbstractWebsite

The diheme cytochrome c peroxidase from Paracoccus denitrificans was modified with the histidine-specific reagent diethyl pyrocarbonate. At low excess of reagent, 1 mol of histidine was modified in the oxidized enzyme, and modification was associated with loss of the ability to form the active state. With time, the modification reversed, and the ability to form the active state was recovered. The agreement between the spectrophotometric measurement of histidine modification and radioactive incorporation using a radiolabeled reagent indicated little modification of other amino acids. However, the reversal of histidine modification observed spectrophotometrically was not matched by loss of radioactivity, and we propose a slow transfer of the ethoxyformyl group to an unidentified amino acid. The presence of CN- bound to the active peroxidatic site of the enzyme led to complete protection of the essential histidine from modification. Limited subtilisin treatment of the native enzyme followed by tryptic digest of the C-terminal fragment (residues 251-338) showed that radioactivity was located in a peptide containing a single histidine at position 275. We propose that this conserved residue, in a highly conserved region, is central to the function of the active mixed-valence state.

Moniz, A, Dinis M.  1996.  Study of Instruments and Tools to Anticipate the Effects of Industrial Change - Portuguese report, Mar. , Number 6604: University Library of Munich, Germany Abstract

This study was produced for the “Study of Instruments and Tools to anticipate the effects of industrial change on employment, trades and vocational qualifications” and for DG V (Employment) of the European Commission in the late 1994. It started when the previous Portuguese government was still ruling, the main policies were defined, and the available instruments were not used in a minimum extend. The new Government, issued from the 1995 elections, proposed “employment” as a major objective with horizontal responsibility. That’s also why there is now a Ministry for Qualifications and Employment, and another one for Solidarity and Social Affairs, not one for Employment and Social Affairs as the previous Government had. But more than that, this objective is considered to need a coordinated and consistent action that involves external affairs, industrial and regional policies, and the policies on education, training and employment, among others. The promotion of the “quality of employment” is being recently done at the working conditions, remuneration, social protection, occupational promotion levels, and the equality of opportunities towards employment and vocational training levels, and finally, the levels of qualification of human resources for a better labour market, education policy and training policy developments. In Portugal, the influence of the industrial change is produced in a top-down way; with (in some cases) an ex post analysis process to formulated training needs. This means that the industrial change impact is produced (normally, unexpectedly), and afterwards the responsible at the company level tries to know which training needs should be formulated in order those effects could be the smoother possible. The training needs at the company level is not based on anticipatory studies, neither is done any long term forecast on qualification, or even employment level.

Moniz, A, Dinis M.  1996.  {Study of Instruments and Tools to Anticipate the Effects of Industrial Change - Portuguese report}, Mar. , Number 6604: University Library of Munich, Germany Abstract

This study was produced for the “Study of Instruments and Tools to anticipate the effects of industrial change on employment, trades and vocational qualifications” and for DG V (Employment) of the European Commission in the late 1994. It started when the previous Portuguese government was still ruling, the main policies were defined, and the available instruments were not used in a minimum extend. The new Government, issued from the 1995 elections, proposed “employment” as a major objective with horizontal responsibility. That’s also why there is now a Ministry for Qualifications and Employment, and another one for Solidarity and Social Affairs, not one for Employment and Social Affairs as the previous Government had. But more than that, this objective is considered to need a coordinated and consistent action that involves external affairs, industrial and regional policies, and the policies on education, training and employment, among others. The promotion of the “quality of employment” is being recently done at the working conditions, remuneration, social protection, occupational promotion levels, and the equality of opportunities towards employment and vocational training levels, and finally, the levels of qualification of human resources for a better labour market, education policy and training policy developments. In Portugal, the influence of the industrial change is produced in a top-down way; with (in some cases) an ex post analysis process to formulated training needs. This means that the industrial change impact is produced (normally, unexpectedly), and afterwards the responsible at the company level tries to know which training needs should be formulated in order those effects could be the smoother possible. The training needs at the company level is not based on anticipatory studies, neither is done any long term forecast on qualification, or even employment level.

Moniz, A.  1996.  Novos modelos de produ{\c c}ão na indústria automóvel? Análise de uma fábrica de motores em Portugal[New production models in the automotive industry? Analysis of an engine factory in Portugal], Jun , Number 7207: University Library of Munich, Germany Abstract

This paper is based in a report for the international project “GERPISA-Labour Relations” on the concept of leader-factory, and in other papers written after that using the case study of the Renault factory in the Aveiro region. This case study is articulated with other studies on engine factories of the same company in Spain, France and Mexico. That study has been co-ordinated by Prof. Juan José Castillo (Univ. Complutense Madrid, Spain). Here we present the results of this empirical research from where were developed the first indicators of a discussion on the concept of new production models and leader-factory that have been studied by the GERPISA international network.

Coelho, AV, Matias PM, Carrondo MA, Tavares P, Moura JJ, Moura I, Fulop V, Hajdu J, Legall J.  1996.  Preliminary crystallographic analysis of the oxidized form of a two mono-nuclear iron centres protein from Desulfovibrio desulfuricans ATCC 27774, Jun. Protein Sci. 5:1189-91., Number 6 AbstractWebsite

Crystals of the fully oxidized form of desulfoferrodoxin were obtained by vapor diffusion from a solution containing 20% PEG 4000, 0.1 M HEPES buffer, pH 7.5, and 0.2 M CaCl2. Trigonal and/or rectangular prisms could be obtained, depending on the temperature used for the crystal growth. Trigonal prisms belong to the rhombohedral space group R32, with a = 112.5 A and c = 63.2 A; rectangular prisms belong to the monoclinic space group C2, with a = 77.7 A, b = 80.9 A, c = 53.9 A, and beta = 98.1 degrees. The crystallographic asymmetric unit of the rhombohedral crystal form contains one molecule. There are two molecules in the asymmetric unit of the monoclinic form, in agreement with the self-rotation function.

Moniz, A.  1996.  {Novos modelos de produção na indústria automóvel? Análise de uma fábrica de motores em Portugal[New production models in the automotive industry? Analysis of an engine factory in Portugal]}, Jun , Number 7207: University Library of Munich, Germany Abstract

This paper is based in a report for the international project “GERPISA-Labour Relations” on the concept of leader-factory, and in other papers written after that using the case study of the Renault factory in the Aveiro region. This case study is articulated with other studies on engine factories of the same company in Spain, France and Mexico. That study has been co-ordinated by Prof. Juan José Castillo (Univ. Complutense Madrid, Spain). Here we present the results of this empirical research from where were developed the first indicators of a discussion on the concept of new production models and leader-factory that have been studied by the GERPISA international network.

Romero, A, Caldeira J, Legall J, Moura I, Moura JJ, Romao MJ.  1996.  Crystal structure of flavodoxin from Desulfovibrio desulfuricans ATCC 27774 in two oxidation states, Jul 1. Eur J Biochem. 239:190-6., Number 1 AbstractWebsite

The crystal structures of the flavodoxin from Desulfovibrio desulfuricans ATCC 27774 have been determined and refined for both oxidized and semi-reduced forms to final crystallographic R-factors of 17.9% (0.8-0.205-nm resolution) and 19.4% (0.8-0.215-nm resolution) respectively. Native flavodoxin crystals were grown from ammonium sulfate with cell constants a = b = 9.59 nm, c=3.37nm (oxidized crystals) and they belong to space group P3(2)21. Semireduced crystals showed some changes in cell dimensions: a = b = 9.51 nm, c=3.35 nm. The three-dimensional structures are similar to other known flavodoxins and deviations are found essentially in the isoalloxazine ring environment. Conformational changes are observed between both redox states and a flip of the Gly61-Met62 peptide bond occurs upon one-electron reduction of the FMN group. These changes influence the redox potential of the oxidized/semiquinone couple. Modulation of the redox potentials is known to be related to the association constant of the FMN group to the protein. The flavodoxin from D. desulfuricans now studied has a large span between E2 (oxidized --> semiquinone) and E1 (semiquinone --> hydroquinone) redox potentials, both these values being substantially more positive within known flavodoxins. A comparison of their FMN environment was made in both oxidation states in order to correlate functional and structural differences.

Coelho, AV, Matias PM, Carrondo MA, Tavares P, Moura JJG, Moura I, Fulop V, Hajdu J, Legall J.  1996.  Preliminary crystallographic analysis of the oxidized form of a two mono-nuclear iron centres protein from Desulfovibrio desulfuricans ATCC 27774, Jul. PROTEIN SCIENCE. {5}:{1189-1191}., Number {6} Abstract

Crystals of the fully oxidized form of desulfoferrodoxin were obtained by vapor diffusion from a solution containing 20% PEG 4000, 0.1 M HEPES buffer, pH 7.5, and 0.2 M CaCl2. Trigonal and/or rectangular prisms could be obtained, depending on the temperature used for the crystal growth. Trigonal prisms belong to the rhombohedral space group R32, with a = 112.5 Angstrom and c = 63.2 Angstrom; rectangular prisms belong to the monoclinic space group C2, with a = 77.7 Angstrom, b = 80.9 Angstrom, c = 53.9 Angstrom, and beta = 98.1 degrees. The crystallographic asymmetric unit of the rhombohedral crystal form contains one molecule. There are two molecules in the asymmetric unit of the monoclinic form, in agreement with the self-rotation function.

Moniz, A, Oliveira P, Bento S.  1996.  Hibrida{\c c}ão de um sistema flex{\'ıvel de produ{\c c}ão: possibilidades de aplica{\c c}ão do conceito de antropocentrismo[Hybridation of a flexible production system: possibilities for an application of t, Feb. , Number 7193: University Library of Munich, Germany Abstract

Today one can understand the wider use of the anthropocentrism concept applied to the production architectures, emerging a new value of the intuitive capacities and human knowledge in the optimization and flexibilization of the manufacturing processes. Having a flexible production and assembly system architecture that exists at UNINOVA-CRI, we will try to develop some exploratory hypothesis on the applicability of the hybridizing concept and its repercussions in the definition of working places, in their organization and formation of working teams. We will underline some aspects that should be taken into consideration when are design such systems, including some ergonomical aspects.

Moniz, A, Oliveira P, Bento S.  1996.  {Hibridação de um sistema flexível de produção: possibilidades de aplicação do conceito de antropocentrismo[Hybridation of a flexible production system: possibilities for an application of the anth}, Feb. , Number 7193: University Library of Munich, Germany Abstract

Today one can understand the wider use of the anthropocentrism concept applied to the production architectures, emerging a new value of the intuitive capacities and human knowledge in the optimization and flexibilization of the manufacturing processes. Having a flexible production and assembly system architecture that exists at UNINOVA-CRI, we will try to develop some exploratory hypothesis on the applicability of the hybridizing concept and its repercussions in the definition of working places, in their organization and formation of working teams. We will underline some aspects that should be taken into consideration when are design such systems, including some ergonomical aspects.

Macedo, AL, Besson S, Moreno C, Fauque G, Moura JJ, Moura I.  1996.  Characterization of a 7Fe ferredoxin isolated from the marine denitrifier Pseudomonas nautica strain 617: spectroscopic and electrochemical studies, Dec 13. Biochem Biophys Res Commun. 229:524-30., Number 2 AbstractWebsite

A 7Fe ferredoxin, isolated from the marine denitrifier Pseudomonas nautica strain 617, was characterized. The NH2-terminal sequence analysis, performed until residue number 56, shows a high similarity with the 7Fe ferredoxins isolated from Azotobacter vinelandii, Pseudomonas putida, and Pseudomonas stutzeri. EPR and NMR spectroscopies identify the presence of both [3Fe-4S] and [4Fe-4S] clusters, with cysteinyl coordination. The electrochemical studies on [Fe-S] clusters show that a fast diffusion-dominated electron transfer, promoted by Mg(II), takes place between the ferredoxin and the glassy carbon electrode. Square wave voltammetry studies gave access to the electrosynthesis of a 4Fe center formed within the [3Fe-4S] core. The [3Fe-4S] cluster exhibited two reduction potentials at -175 and -680 +/- 10 mV and the [4Fe-4S] cluster was characterized by an unusually low reduction potential of -715 +/- 10 mV, at pH 7.6

Huber, R, Hof P, Duarte RO, Moura JJ, Moura I, Liu MY, Legall J, Hille R, Archer M, Romao MJ.  1996.  A structure-based catalytic mechanism for the xanthine oxidase family of molybdenum enzymes, Aug 20. Proc Natl Acad Sci U S A. 93:8846-51., Number 17 AbstractWebsite

The crystal structure of the xanthine oxidase-related molybdenum-iron protein aldehyde oxido-reductase from the sulfate reducing anaerobic Gram-negative bacterium Desulfovibrio gigas (Mop) was analyzed in its desulfo-, sulfo-, oxidized, reduced, and alcohol-bound forms at 1.8-A resolution. In the sulfo-form the molybdenum molybdopterin cytosine dinucleotide cofactor has a dithiolene-bound fac-[Mo, = O, = S, ---(OH2)] substructure. Bound inhibitory isopropanol in the inner compartment of the substrate binding tunnel is a model for the Michaelis complex of the reaction with aldehydes (H-C = O,-R). The reaction is proposed to proceed by transfer of the molybdenum-bound water molecule as OH- after proton transfer to Glu-869 to the carbonyl carbon of the substrate in concert with hydride transfer to the sulfido group to generate [MoIV, = O, -SH, ---(O-C = O, -R)). Dissociation of the carboxylic acid product may be facilitated by transient binding of Glu-869 to the molybdenum. The metal-bound water is replenished from a chain of internal water molecules. A second alcohol binding site in the spacious outer compartment may cause the strong substrate inhibition observed. This compartment is the putative binding site of large inhibitors of xanthine oxidase.

Caldeira, J, Feicht R, White H, Teixeira M, Moura JJ, Simon H, Moura I.  1996.  EPR and Mossbauer spectroscopic studies on enoate reductase, Aug 2. J Biol Chem. 271:18743-8., Number 31 AbstractWebsite

Enoate reductase (EC 1.3.1.31) is a protein isolated from Clostridium tyrobutyricum that contains iron, labile sulfide, FAD, and FMN. The enzyme reduces the alpha,beta carbon-carbon double bond of nonactivated 2-enoates and in a reversible way that of 2-enals at the expense of NADH or reduced methyl viologen. UV-visible and EPR potentiometric titrations detect a semiquinone species in redox intermediate states characterized by an isotropic EPR signal at g = 2.0 without contribution at 580 nm. EPR redox titration shows two widely spread mid-point redox potentials (-190 and -350 mV at pH 7. 0), and a nearly stoichiometric amount of this species is detected. The data suggest the semiquinone radical has an anionic nature. In the reduced form, the [Fe-S] moiety is characterized by a single rhombic EPR spectrum, observed in a wide range of temperatures (4. 2-60 K) with g values at 2.013, 1.943, and 1.860 (-180 mV at pH 7.0). The gmax value is low when compared with what has been reported for other iron-sulfur clusters. Mossbauer studies reveal the presence of a [4Fe-4S]+2/+1 center. One of the subcomponents of the spectrum shows an unusually large value of quadrupole splitting (ferrous character) in both the oxidized and reduced states. Substrate binding to the reduced enzyme induces subtle changes in the spectroscopic Mossbauer parameters. The Mossbauer data together with known kinetic information suggest the involvement of this iron-sulfur center in the enzyme mechanism.

Goodfellow, BJ, Tavares P, Romao MJ, Czaja C, Rusnak F, Legall J, Moura I, Moura JJG.  1996.  The solution structure of desulforedoxin, a simple iron-sulfur protein - An NMR study of the zinc derivative, Aug. Journal of Biological Inorganic Chemistry. 1:341-354., Number 4 AbstractWebsite

Desulforedoxin is a simple dimeric protein isolated from Desulfovibrio gigas containing a distorted rubredoxin-like center with one iron coordinated by four cysteinyl residues (7.9 kDa with a 36-amino-acid monomer). H-1 NMR spectra of the oxidized Dx(Fe3+) and reduced Dx(Fe2+) forms were analyzed. The spectra show substantial line broadening due to the paramagnetism of iron. However, very low-field-shifted resonances, assigned to H beta protons, were observed in the reduced state and their temperature dependence analyzed. The active site of Dx was reconstituted with zinc, and its solution structure was determined using 2D NMR methods. This diamagnetic form gave high-resolution NMR data enabling the identification of all the amino acid spin systems. Sequential assignment and the determination of secondary structural elements was attempted using 2D NOESY experiments. However, because of the symmetrical dimer nature of the protein standard, NMR sequential assignment methods could not resolve all cross peaks due to inter- and intra-chain effects. The X-ray structure enabled the spatial relationship between the monomers to be obtained, and resolved the assignment problems. Secondary structural features could be identified from the NMR data; an antiparallel beta-sheet running from D5 to V18 with a well-defined beta-turn around cysteines C9 and C12. The section G22 to T25 is poorly defined by the NMR data and is followed by a turn around V27-C29. The C-terminus ends up near residues V6 and Y7. Distance geometry (DG) calculations allowed families of structures to be generated from the NMR data. A family of structures with a low target function violation for the Dr monomer and dimer were found to have secondary structural elements identical to those seen in the X-ray structure. The amide protons for G4, D5, G13, L11 NH and Q14 NH epsilon amide protons, H-bonded in the X-ray structure, were not seen by NMR as slowly exchanging, while structural disorder at the N-terminus, for the backbone at E10 and for the section G22-T25, was observed. Comparison between the Fe and Zn forms of Dr suggests that metal substitution does not have an effect on the structure of the protein.

Goodfellow, BJ, Tavares P, Romão MJ, Czaja C, Rusnak F, Legall J, Moura I, Moura JJG.  1996.  The solution structure of desulforedoxin, a simple iron-sulfur protein - An NMR study of the zinc derivative, Aug. JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY. {1}:{341-354}., Number {4} Abstract

Desulforedoxin is a simple dimeric protein isolated from Desulfovibrio gigas containing a distorted rubredoxin-like center with one iron coordinated by four cysteinyl residues (7.9 kDa with a 36-amino-acid monomer). H-1 NMR spectra of the oxidized Dx(Fe3+) and reduced Dx(Fe2+) forms were analyzed. The spectra show substantial line broadening due to the paramagnetism of iron. However, very low-field-shifted resonances, assigned to H beta protons, were observed in the reduced state and their temperature dependence analyzed. The active site of Dx was reconstituted with zinc, and its solution structure was determined using 2D NMR methods. This diamagnetic form gave high-resolution NMR data enabling the identification of all the amino acid spin systems. Sequential assignment and the determination of secondary structural elements was attempted using 2D NOESY experiments. However, because of the symmetrical dimer nature of the protein standard, NMR sequential assignment methods could not resolve all cross peaks due to inter- and intra-chain effects. The X-ray structure enabled the spatial relationship between the monomers to be obtained, and resolved the assignment problems. Secondary structural features could be identified from the NMR data; an antiparallel beta-sheet running from D5 to V18 with a well-defined beta-turn around cysteines C9 and C12. The section G22 to T25 is poorly defined by the NMR data and is followed by a turn around V27-C29. The C-terminus ends up near residues V6 and Y7. Distance geometry (DG) calculations allowed families of structures to be generated from the NMR data. A family of structures with a low target function violation for the Dr monomer and dimer were found to have secondary structural elements identical to those seen in the X-ray structure. The amide protons for G4, D5, G13, L11 NH and Q14 NH epsilon amide protons, H-bonded in the X-ray structure, were not seen by NMR as slowly exchanging, while structural disorder at the N-terminus, for the backbone at E10 and for the section G22-T25, was observed. Comparison between the Fe and Zn forms of Dr suggests that metal substitution does not have an effect on the structure of the protein.

Pereira, AS, Franco R, Feio MJ, Pinto C, Lampreia J, Reis MA, Calvete J, Moura I, Beech I, Lino AR, Moura JJ.  1996.  Characterization of representative enzymes from a sulfate reducing bacterium implicated in the corrosion of steel, Apr 16. Biochem Biophys Res Commun. 221:414-21., Number 2 AbstractWebsite

This communication reports the isolation, purification and characterization of key enzymes involved in dissimilatory sulfate reduction of a sulfate reducing bacterium classified as Desulfovibrio desulfuricans subspecies desulfuricans New Jersey (NCIMB 8313) (Ddd NJ). The chosen strain, originally recovered from a corroding cast iron heat exchanger, was grown in large scale batch cultures. Physico-chemical and spectroscopic studies of the purified enzymes were carried out. These analyses revealed a high degree of similarity between proteins isolated from the DddNJ strain and the homologous proteins obtained from Desulfomicrobium baculatus Norway 4. In view of the results obtained, taxonomic reclassification of Desulfovibrio desulfuricans subspecies desulfuricans New Jersey (NCIMB 8313) into Desulfomicrobium baculatus (New Jersey) is proposed.

Moura, JJG, Goodfellow BJ, Romao MJ, Rusnak F, Moura I.  1996.  Analysis, design and engineering of simple iron-sulfur proteins: Tales from rubredoxin and desulforedoxin, 1996. Comments on Inorganic Chemistry. 19:47-+., Number 1 AbstractWebsite

The most thoroughly characterized non-heme iron center in biology is Rubredoxin, the simplest member of the iron-sulfur: class of metalloproteins. Rubredoxin contains a high-spin iron atom with tetrahedral coordination by four cysteinyl sulfur atoms. A structural variant of this center is found in Desulforedoxin, the smallest known Rubredoxin type protein. The 3D structure of both Rd and Dr has been determined at high resolution. These two proteins can therefore be used as case studies in which structural control by the polypeptide chain over the metal site can be discussed in detail.

Pina, F, Benedito L, Melo MJ, Parola AJ, Bernardo A.  1996.  Photochemistry of 3,4'-dimethoxy-7-hydroxyflavylium chloride - Photochromism and excited-state proton transfer, 1996. Journal of the Chemical Society-Faraday Transactions. 92:1693-1699. AbstractWebsite

The synthetic compound 3,4'-dimethoxy-7-hydroxyflavylium chloride gives rise, in aqueous solution at moderately acidic pH, to a pH-dependent equilibrium between the flavylium cation, hemiacetal, (Z)-chalcone and a small amount of quinonoidal base. The distribution, as a function of pH, of the molar fractions of the several species present in solution have been calculated on the basis of H-1 NMR and pH jump experiments monitored by stopped-flow and conventional UV-VIS spectrophotometry, and highperformance liquid chromatography (HPLC). The compound shows interesting photochemical properties: (i) at pH 4.0 it presents a photochromic effect that converts (Z)-chalcone into hemiacetal, the reaction being reversible in the dark and (ii) excited-state proton transfer is observed between the flavylium cation and quinonoidal base. An appropriate formalism to quantify the experimental results has been developed. The formalism allows determination of the pH-dependent molar fraction distribution of the several anthocyanin forms present at equilibrium, as well as predicting the distribution of the molar fractions prior to equilibrium.

Parola, AJ, Pina F, Ferreira E, Maestri M, Balzani V.  1996.  Photoinduced electron- and energy-transfer processes of biacetyl imprisoned in a hemicarcerand, 1996. Journal of the American Chemical Society. 118:11610-11616. AbstractWebsite

The energy- and electron-transfer quenching processes of the lowest triplet excited state of biacetyl (2,3-butanedione) imprisoned in a hemicarcerand have been systematically investigated in CH2Cl2 solution at room temperature. Twenty potential quenchers have been used, including ten triplet energy accepters (mostly, aromatic hydrocarbons) and nine electron donors (mostly, aromatic amines). Bimolecular rate constants for the quenching processes were obtained by Stern-Volmer analysis and compared with those found for the quenching of free biacetyl. In the electron-transfer processes, aromatic amines with oxidation potential from +0.015 V (N,N,N',N'-tetramethyl-p-phenylenediamine) to +0.83 V (diphenylamine) quench free biacetyl at the diffusion-controlled limit, whereas for imprisoned biacetyl the rate constant decreases (roughly in a linear manner) from 4.0 x 10(8) to 1.2 x 10(5) M(-1) s(-1) As far as energy-transfer is concerned, the rate constant for the quenching of free biacetyl increases with decreasing Delta G degrees and reaches the diffusion-controlled plateau value (k(q) similar to 10(10) M(-1) s(-1)) for Delta G degrees similar to 0.1 eV, whereas for imprisoned biacetyl a scattered, bell-shaped log k(q) vs Delta G degrees plot is obtained, with a maximum value (similar to 10(6) M(-1) s(-1)) much below the diffusion-controlled limit. The results obtained show that the walls of the hemicarcerand allow only very weak electronic interaction between incarcerated triplet biacetyl and external quenchers. A brief discussion of the results obtained in the light of current energy- and electron-transfer theories is presented.

Moura, JJG, Goodfellow BJ, Romao MJ, Rusnak F, Moura I.  1996.  Analysis, design and engineering of simple iron-sulfur proteins: Tales from rubredoxin and desulforedoxin. Comments on Inorganic Chemistry. 19:47-+., Number 1 AbstractWebsite
n/a
Pereira, AS, Franco R, Feio MJ, Pinto C, Lampreia J, Reis MA, Calvete J, Moura I, Beech I, Lino AR, Moura JJG.  1996.  Characterization of representative enzymes from a sulfate reducing bacterium implicated in the corrosion of steel. Biochemical And Biophysical Research Communications. {221}:{414-421}., Number {2}, 525 B ST, STE 1900, SAN DIEGO, CA 92101-4495: ACADEMIC PRESS INC JNL-COMP SUBSCRIPTIONS Abstract

This communication reports the isolation, purification and characterization of key enzymes involved in dissimilatory sulfate reduction of a sulfate reducing bacterium classified as Desulfovibrio desulfuricans subspecies desulfuricans New Jersey (NCIMB 8313) (Ddd NJ). The chosen strain, originally recovered from a corroding cast iron heat exchanger, was grown in large scale batch cultures. Physico-chemical and spectroscopic studies of the purified enzymes were carried out. These analyses revealed a high degree of similarity between proteins isolated from the DddNJ strain and the homologous proteins obtained from Desulfomicrobium baculatus Norway 4. In view of the results obtained, taxonomic reclassification of Desulfovibrio desulfuricans subspecies desulfuricans New Jersey (NCIMB 8313) into Desulfomicrobium baculatus (New Jersey) is proposed. (C) 1996 Academic Press, Inc.

Romero, A, Caldeira J, Legall J, Moura I, Moura JJG, Romao MJ.  1996.  Crystal structure of flavodoxin from Desulfovibrio desulfuricans ATCC 27774 in two oxidation states. European Journal of Biochemistry. 239:190-196., Number 1 AbstractWebsite
n/a
loading