Publications

Export 153 results:
Sort by: Author [ Title  (Asc)] Type Year
A B C D E F G H I J K L M N O P Q [R] S T U V W X Y Z   [Show ALL]
1
The 1.4 angstrom resolution structure of Paracoccus pantotrophus pseudoazurin, Najmudin, Shabir, Pauleta Sofia R., Moura Isabel, and Romao Maria J. , Acta Crystallographica Section F-Structural Biology and Crystallization Communications, Jun, Volume 66, p.627-635, (2010) AbstractWebsite

Pseudoazurins are small type 1 copper proteins that are involved in the flow of electrons between various electron donors and acceptors in the bacterial periplasm, mostly under denitrifying conditions. The previously determined structure of Paracoccus pantotrophus pseudoazurin in the oxidized form was improved to a nominal resolution of 1.4 angstrom, with R and R(free) values of 0.188 and 0.206, respectively. This high-resolution structure makes it possible to analyze the interactions between the monomers and the solvent structure in detail. Analysis of the high-resolution structure revealed the structural regions that are responsible for monomer-monomer recognition during dimer formation and for protein-protein interaction and that are important for partner recognition. The pseudoazurin structure was compared with other structures of various type 1 copper proteins and these were grouped into families according to similarities in their secondary structure; this may be useful in the annotation of copper proteins in newly sequenced genomes and in the identification of novel copper proteins.

A
Actin as a potential target for decavanadate, Ramos, S., Moura J. J., and Aureliano M. , J Inorg Biochem, Dec, Volume 104, Number 12, p.1234-9, (2011) AbstractWebsite

ATP prevents G-actin cysteine oxidation and vanadyl formation specifically induced by decavanadate, suggesting that the oxometalate-protein interaction is affected by the nucleotide. The ATP exchange rate is increased by 2-fold due to the presence of decavanadate when compared with control actin (3.1x10(-3) s(-1)), and an apparent dissociation constant (k(dapp)) of 227.4+/-25.7 muM and 112.3+/-8.7 muM was obtained in absence or presence of 20 muM V(10), respectively. Moreover, concentrations as low as 50 muM of decameric vanadate species (V(10)) increases the relative G-actin intrinsic fluorescence intensity by approximately 80% whereas for a 10-fold concentration of monomeric vanadate (V(1)) no effects were observed. Upon decavanadate titration, it was observed a linear increase in G-actin hydrophobic surface (2.6-fold), while no changes were detected for V(1) (0-200 muM). Taken together, three major ideas arise: i) ATP prevents decavanadate-induced G-actin cysteine oxidation and vanadate reduction; ii) decavanadate promotes actin conformational changes resulting on its inactivation, iii) decavanadate has an effect on actin ATP binding site. Once it is demonstrated that actin is a new potential target for decavanadate, being the ATP binding site a suitable site for decavanadate binding, it is proposed that some of the biological effects of vanadate can be, at least in part, explained by decavanadate interactions with actin.

Amyloid beta-peptide disrupts mitochondrial membrane lipid and protein structure: protective role of tauroursodeoxycholate, Rodrigues, C. M., Sola S., Brito M. A., Brondino C. D., Brites D., and Moura J. J. , Biochem Biophys Res Commun, Feb 23, Volume 281, Number 2, p.468-74, (2001) AbstractWebsite

Mitochondria have been implicated in the cytotoxicity of amyloid beta-peptide (A beta), which accumulates as senile plaques in the brain of Alzheimer's disease patients. Tauroursodeoxycholate (TUDC) modulates cell death, in part, by preventing mitochondrial membrane perturbation. Using electron paramagnetic resonance spectroscopy analysis of isolated mitochondria, we tested the hypothesis that A beta acts locally in mitochondrial membranes to induce oxidative injury, leading to increased membrane permeability and subsequent release of caspase-activating factors. Further, we intended to determine the role of TUDC at preventing A beta-induced mitochondrial membrane dysfunction. The results demonstrate oxidative injury of mitochondrial membranes during exposure to A beta and reveal profound structural changes, including modified membrane lipid polarity and disrupted protein mobility. Cytochrome c is released from the intermembrane space of mitochondria as a consequence of increased membrane permeability. TUDC, but not cyclosporine A, almost completely abrogated A beta-induced perturbation of mitochondrial membrane structure. We conclude that A beta directly induces cytochrome c release from mitochondria through a mechanism that is accompanied by profound effects on mitochondrial membrane redox status, lipid polarity, and protein order. TUDC can directly suppress A beta-induced disruption of the mitochondrial membrane structure, suggesting a neuroprotective role for this bile salt.

Analysis of the activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase through an electron transfer chain, Paes de Sousa, P. M., Rodrigues D., Timoteo C. G., Simoes Goncalves M. L., Pettigrew G. W., Moura I., Moura J. J., and Correia dos Santos M. M. , J Biol Inorg Chem, Aug, Volume 16, Number 6, p.881-8, (2011) AbstractWebsite

The activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase (CCP) was probed through the mediated electrochemical catalysis by its physiological electron donor, P. stutzeri cytochrome c-551. A comparative study was carried out, by performing assays with the enzyme in the resting oxidized state as well as in the mixed-valence activated form, using cyclic voltammetry and a pyrolytic graphite membrane electrode. In the presence of both the enzyme and hydrogen peroxide, the peak-like signal of cytochrome c-551 is converted into a sigmoidal wave form characteristic of an E(r)C'(i) catalytic mechanism. An intermolecular electron transfer rate constant of (4 +/- 1) x 10(5) M(-1) s(-1) was estimated for both forms of the enzyme, as well as a similar Michaelis-Menten constant. These results show that neither the intermolecular electron transfer nor the catalytic activity is kinetically controlled by the activation mechanism of CCP in the case of the P. stutzeri enzyme. Direct enzyme catalysis using protein film voltammetry was unsuccessful for the analysis of the activation mechanism, since P. stutzeri CCP undergoes an undesirable interaction with the pyrolytic graphite surface. This interaction, previously reported for the Paracoccus pantotrophus CCP, induces the formation of a non-native conformation state of the electron-transferring haem, which has a redox potential 200 mV lower than that of the native state and maintains peroxidatic activity.

Analysis, design and engineering of simple iron-sulfur proteins: Tales from rubredoxin and desulforedoxin, Moura, J. J. G., Goodfellow B. J., Romao M. J., Rusnak F., and Moura I. , Comments on Inorganic Chemistry, 1996, Volume 19, Number 1, p.47-+, (1996) 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.

Aromatic aldehydes at the active site of aldehyde oxidoreductase from Desulfovibrio gigas: reactivity and molecular details of the enzyme-substrate and enzyme-product interaction, Correia, H., Marangon J., Brondino C. D., Moura J. J. G., Romao M. J., Gonzalez P. J., and Santos-Silva T. , J Biol Inorg Chem, Volume 20, p.219-229, (2015)
Association of Zn, Cu, Cd and Pb with protein fractions and sub-cellular partitioning in the digestive gland of Octopus vulgaris living in habitats with different metal levels, Raimundo, J., Vale C., Duarte R., and Moura I. , Chemosphere, Nov, Volume 81, Number 10, p.1314-1319, (2010) AbstractWebsite

Zinc Cu Cd and Pb concentrations were determined in protein fractions of digestive gland and in the whole digestive gland of Octopus vulgaris collected from two areas of the Portuguese coast Approximately 95% of Zn 99% of Cu 85-96% of Cd and 77-86% of Pb were stored in the cytosol suggesting the predominance of cytosolic proteins in the trapping these elements Gel filtration chromatography evidenced the presence of two major groups of proteins with high molecular weight (HMW 144 000-130 000 Da) and low molecular weight (LMW 11 000-6000 Da) The following metal-protein associations were found Zn was distributed between HMW and LMW Cu and Cd in LMW proteins with a minor association with HMW and Pb in HMW proteins The strong positive correlations between Cd Zn and Cu and LMW proteins point to the presence of metalloproteins with high affinity to these elements A shift was registered between the maximum of the ratio 254 280 nm and metal concentrations in the chromatographic profiles This shift may result from metallothioneins having a small participation in the metal binding or protein purification was insufficient and various LMW proteins may be interfering (C) 2010 Elsevier Ltd All rights reserved

B
Benefits of membrane electrodes in the electrochemistry of metalloproteins: mediated catalysis of Paracoccus pantotrophus cytochrome c peroxidase by horse cytochrome c: a case study, Paes de Sousa, P. M., Pauleta S. R., Rodrigues D., Simoes Goncalves M. L., Pettigrew G. W., Moura I., Moura J. J., and Correia dos Santos M. M. , J Biol Inorg Chem, Jun, Volume 13, Number 5, p.779-87, (2008) AbstractWebsite

A comparative study of direct and mediated electrochemistry of metalloproteins in bulk and membrane-entrapped solutions is presented. This work reports the first electrochemical study of the electron transfer between a bacterial cytochrome c peroxidase and horse heart cytochrome c. The mediated catalysis of the peroxidase was analysed both using the membrane electrode configuration and with all proteins in solution. An apparent Michaelis constant of 66 +/- 4 and 42 +/- 5 microM was determined at pH 7.0 and 0 M NaCl for membrane and bulk solutions, respectively. The data revealed that maximum activity occurs at 50 mM NaCl, pH 7.0, with intermolecular rate constants of (4.4 +/- 0.5) x 10(6) and (1.0 +/- 0.5) x 10(6) M(-1) s(-1) for membrane-entrapped and bulk solutions, respectively. The influence of parameters such as pH or ionic strength on the mediated catalytic activity was analysed using this approach, drawing attention to the fact that careful analysis of the results is needed to ensure that no artefacts are introduced by the use of the membrane configuration and/or promoters, and therefore the dependence truly reflects the influence of these parameters on the (mediated) catalysis. From the pH dependence, a pK of 7.5 was estimated for the mediated enzymatic catalysis.

Bilirubin directly disrupts membrane lipid polarity and fluidity, protein order, and redox status in rat mitochondria, Rodrigues, C. M., Sola S., Brito M. A., Brites D., and Moura J. J. , J Hepatol, Mar, Volume 36, Number 3, p.335-41, (2002) AbstractWebsite

BACKGROUND/AIMS: Unconjugated bilirubin (UCB) impairs crucial aspects of cell function and induces apoptosis in primary cultured neurones. While mechanisms of cytotoxicity begin to unfold, mitochondria appear as potential primary targets. METHODS: We used electron paramagnetic resonance spectroscopy analysis of isolated rat mitochondria to test the hypothesis that UCB physically interacts with mitochondria to induce structural membrane perturbation, leading to increased permeability, and subsequent release of apoptotic factors. RESULTS: Our data demonstrate profound changes on mitochondrial membrane properties during incubation with UCB, including modified membrane lipid polarity and fluidity (P<0.01), as well as disrupted protein mobility (P<0.001). Consistent with increased permeability, cytochrome c was released from the intermembrane space (P<0.01), perhaps uncoupling the respiratory chain and further increasing oxidative stress (P<0.01). Both ursodeoxycholate, a mitochondrial-membrane stabilising agent, and cyclosporine A, an inhibitor of the permeability transition, almost completely abrogated UCB-induced perturbation. CONCLUSIONS: UCB directly interacts with mitochondria influencing membrane lipid and protein properties, redox status, and cytochrome c content. Thus, apoptosis induced by UCB may be mediated, at least in part, by physical perturbation of the mitochondrial membrane. These novel findings should ultimately prove useful to our evolving understanding of UCB cytotoxicity.

Biochemical and spectroscopic characterization of an aldehyde oxidoreductase isolated from Desulfovibrio aminophilus, Thapper, A., Rivas M. G., Brondino C. D., Ollivier B., Fauque G., Moura I., and Moura J. J. , J Inorg Biochem, Jan, Volume 100, Number 1, p.44-50, (2006) AbstractWebsite

Aldehyde oxidoreductase (AOR) activity has been found in a number of sulfate-reducing bacteria. The enzyme that is responsible for the conversion of aldehydes to carboxylic acids is a mononuclear molybdenum enzyme belonging to the xanthine oxidase family. We report here the purification and characterization of AOR isolated from the sulfate-reducing bacterium Desulfovibrio (D.) aminophilus DSM 12254, an aminolytic strain performing thiosulfate dismutation. The enzyme is a homodimer (ca. 200 kDa), containing a molybdenum centre and two [2Fe-2S] clusters per monomer. UV/Visible and electron paramagnetic resonance (EPR) spectra of D. aminophilus AOR recorded in as-prepared and reduced states are similar to those obtained in AORs from Desulfovibrio gigas, Desulfovibrio desulfuricans and Desulfovibrio alaskensis. Despite AOR from D. aminophilus is closely related to other AORs, it presents lower activity towards aldehydes and no activity towards N-heterocyclic compounds, which suggests another possible role for this enzyme in vivo. A comparison of the molecular and EPR properties of AORs from different Desulfovibrio species is also included.

Biochemical and spectroscopic characterization of overexpressed fuscoredoxin from Escherichia coli, Pereira, A. S., Tavares P., Krebs C., Huynh B. H., Rusnak F., Moura I., and Moura J. J. , Biochem Biophys Res Commun, Jun 24, Volume 260, Number 1, p.209-15, (1999) AbstractWebsite

Fuscoredoxin is a unique iron containing protein of yet unknown function originally discovered in the sulfate reducers of the genus Desulfovibrio. It contains two iron-sulfur clusters: a cubane [4Fe-4S] and a mixed oxo- and sulfido-bridged 4Fe cluster of unprecedented structure. The recent determination of the genomic sequence of Escherichia coli (E. coli) has revealed a homologue of fuscoredoxin in this facultative microbe. The presence of this gene in E. coli raises interesting questions regarding the function of fuscoredoxin and whether this gene represents a structural homologue of the better-characterized Desulfovibrio proteins. In order to explore the latter, an overexpression system for the E. coli fuscoredoxin gene was devised. The gene was cloned from genomic DNA by use of the polymerase chain reaction into the expression vector pT7-7 and overexpressed in E. coli BL21(DE3) cells. After two chromatographic steps a good yield of recombinant protein was obtained (approximately 4 mg of pure protein per liter of culture). The purified protein exhibits an optical spectrum characteristic of the homologue from D. desulfuricans, indicating that cofactor assembly was accomplished. Iron analysis indicated that the protein contains circa 8 iron atoms/molecule which were shown by EPR and Mossbauer spectroscopies to be present as two multinuclear clusters, albeit with slightly altered spectroscopic features. A comparison of the primary sequences of fuscoredoxins is presented and differences on cluster coordination modes are discussed on the light of the spectroscopic data.

Biochemical/spectroscopic characterization and preliminary X-ray analysis of a new aldehyde oxidoreductase isolated from Desulfovibrio desulfuricans ATCC 27774, Duarte, R. O., Archer M., Dias J. M., Bursakov S., Huber R., Moura I., Romao M. J., and Moura J. J. , Biochem Biophys Res Commun, Feb 24, Volume 268, Number 3, p.745-9, (2000) AbstractWebsite

Aldehyde oxidoreductase (AOR) activity has been found in different sulfate reducing organisms (Moura, J. J. G., and Barata, B. A. S. (1994) in Methods in Enzymology (Peck, H. D., Jr., and LeGall, J., Eds.), Vol. 243, Chap. 4. Academic Press; Romao, M. J., Knablein, J., Huber, R., and Moura, J. J. G. (1997) Prog. Biophys. Mol. Biol. 68, 121-144). The enzyme was purified to homogeneity from extracts of Desulfovibrio desulfuricans (Dd) ATCC 27774, a sulfate reducer that can use sulfate or nitrate as terminal respiratory substrates. The protein (AORDd) is described as a homodimer (monomer, circa 100 kDa), contains a Mo-MCD pterin, 2 x [2Fe-2S] clusters, and lacks a flavin group. Visible and EPR spectroscopies indicate a close similarity with the AOR purified from Desulfovibrio gigas (Dg) (Barata, B. A. S., LeGall, J., and Moura, J. J. G. (1993) Biochemistry 32, 11559-11568). Activity and substrate specificity for different aldehydes were determined. EPR studies were performed in native and reduced states of the enzyme and after treatment with ethylene glycol and dithiothreitol. The AORDd was crystallized using ammonium sulfate as precipitant and the crystals belong to the space group P6(1)22, with unit cell dimensions a = b = 156.4 and c = 177.1 A. These crystals diffract to beyond 2.5 A resolution and a full data set was measured on a rotating anode generator. The data were used to solve the structure by Patterson Search methods, using the model of AORDg.

Bioelectricity generation using long-term operated biocathode: RFLP based microbial diversity analysis, Ramanaiaha, S. V., Cordas C. M., Matias S. C., Reddyd M. V., Leitão J. H., and Fonseca L. P. , Biotechnology Reports, Volume 32, p.e00693, (2021)
C
Carbon dioxide utilisation - bioelectrochemical approaches, C.M., Cordas, J.J.G. Moura, A. Escapa, and R. Mateos , Enzymes for Solving Humankind's Problems, Moura J.J.G., Moura I., Maia L.B. (eds), p.83-108, (2021)
Changes in metabolic pathways of Desulfovibrio alaskensis G20 cells induced by molybdate excess, Nair, R. R., Silveira C. M., Diniz M. S., Almeida M. G., Moura J. J. G., and Rivas M. G. , J Biol Inorg Chem, Volume 20, p.311–322, (2015)
Characterization of recombinant Desulfovibrio gigas ferredoxin, Rodrigues, P., Graca F., Macedo A. L., Moura I., and Moura J. J. , Biochem Biophys Res Commun, Nov 30, Volume 289, Number 2, p.630-3, (2001) AbstractWebsite

Dg ferredoxin gene was cloned using the polymerase chain reaction (PCR), inserted into vector pT7-7, and overexpressed in Escherichia coli (E. coli) grown in aerobic media. The recombinant protein is a dimer and contains a [3Fe-4S] cluster per monomer. EPR and (1)H NMR data of recombinant and wild-type protein are compared.

Characterization of representative enzymes from a sulfate reducing bacterium implicated in the corrosion of steel, Pereira, A. S., Franco R., Feio M. J., Pinto C., Lampreia J., Reis M. A., Calvete J., Moura I., Beech I., Lino A. R., and Moura J. J. , Biochem Biophys Res Commun, Apr 16, Volume 221, Number 2, p.414-21, (1996) 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.

Chromatographic-based methods for pesticide determination in honey: An overview, Rial-Otero, R., Gaspar E. M., Moura I., and Capelo J. L. , Talanta, Feb 15, Volume 71, Number 2, p.503-514, (2007) AbstractWebsite

Nowadays the control of pesticides in honey is an issue of primary health importance as consequence of the increasing content of these chemicals in the aforementioned matrix. This poisoning has led to the worldwide increasing loss of bees since 1995. From Europe to Canada, scientist, beekeepers and chemical companies disagree about the reasons that have led to colony losses higher than 50% in some areas. This problem has become a public health issue due to the high honey worldwide consumption. The presence of pesticides in honey has been directly related to bees' mortality by some researchers through pesticide presence in (1) pollen, (2) honeycomb walls, (3) own bees and (4) honey. In this work we describe the actual state-of-the-art for pesticides determination in honey along with a review in this subject focused on sample treatments and instrumentation. Finally, future trends are also commented. (c) 2006 Elsevier B.V. All rights reserved.

Cobalt-, zinc- and iron-bound forms of adenylate kinase (AK) from the sulfate-reducing bacterium Desulfovibrio gigas: purification, crystallization and preliminary X-ray diffraction analysis, Kladova, A. V., Gavel O. Y., Mukhopaadhyay A., Boer D. R., Teixeira S., Shnyrov V. L., Moura I., Moura J. J., Romao M. J., Trincao J., and Bursakov S. A. , Acta Crystallogr Sect F Struct Biol Cryst Commun, Sep 1, Volume 65, Number Pt 9, p.926-9, (2009) AbstractWebsite

Adenylate kinase (AK; ATP:AMP phosphotransferase; EC 2.7.4.3) is involved in the reversible transfer of the terminal phosphate group from ATP to AMP. AKs contribute to the maintenance of a constant level of cellular adenine nucleotides, which is necessary for the energetic metabolism of the cell. Three metal ions, cobalt, zinc and iron(II), have been reported to be present in AKs from some Gram-negative bacteria. Native zinc-containing AK from Desulfovibrio gigas was purified to homogeneity and crystallized. The crystals diffracted to beyond 1.8 A resolution. Furthermore, cobalt- and iron-containing crystal forms of recombinant AK were also obtained and diffracted to 2.0 and 3.0 A resolution, respectively. Zn(2+)-AK and Fe(2+)-AK crystallized in space group I222 with similar unit-cell parameters, whereas Co(2+)-AK crystallized in space group C2; a monomer was present in the asymmetric unit for both the Zn(2+)-AK and Fe(2+)-AK forms and a dimer was present for the Co(2+)-AK form. The structures of the three metal-bound forms of AK will provide new insights into the role and selectivity of the metal in these enzymes.

Comparative electrochemical study of superoxide reductases, Cordas, C. M., Raleiras P., Auchere F., Moura I., and Moura J. J. , Eur Biophys J, Dec 6, Volume 41, Number 12, p.209-215, (2012)
A Comparison between Vanadyl, Vanadate, and Decavanadate Effects in Actin Structure and Function: Combination of Several Spectroscopic Studies, Ramos, S., Moura J. J. G., and Aureliano M. , Spectroscopy: An International Journal, Volume 27, p.355-359, (2012)
The complete catalytic mechanism of Xanthine Oxidase: a computational study, Fernandes, H., Maia L., Ribeiro P. M., J.J.G. Moura, and Cerqueira N. M. , Inorg Chem Front, Volume 8, p.405, (2021)
Construction of effective disposable biosensors for point-of-care testing of nitrite, Monteiro, T., Rodrigues P. R., Gonçalves A. L., Moura J. J. G., Anorga L., Jubete E., Piknova B., Schechter A. N., Silveira C. M., and Almeida M. G. , Talanta, Volume 142, p.246-251, (2015)
Conversion of desulforedoxin into a rubredoxin center, Yu, L., Kennedy M., Czaja C., Tavares P., Moura J. J., Moura I., and Rusnak F. , Biochem Biophys Res Commun, Feb 24, Volume 231, Number 3, p.679-82, (1997) AbstractWebsite

Rubredoxin and desulforedoxin both contain an Fe(S-Cys)4 center. However, the spectroscopic properties of the center in desulforedoxin differ from rubredoxin. These differences arise from a distortion of the metal site hypothesized to result from adjacent cysteine residues in the primary sequence of desulforedoxin. Two desulforedoxin mutants were generated in which either a G or P-V were inserted between adjacent cysteines. Both mutants exhibited optical spectra with maxima at 278, 345, 380, 480, and 560 nm while the low temperature X-band EPR spectra indicated highspin Fe3+ ions with large rhombic distortions (E/D = 0.21-0.23). These spectroscopic properties are distinct from wild type desulforedoxin and virtually identical to rubredoxin.

Copper-substituted forms of the wild type and C42A variant of rubredoxin, Thapper, A., Rizzi A. C., Brondino C. D., Wedd A. G., Pais R. J., Maiti B. K., Moura I., Pauleta S. R., and Moura J. J. G. , J Inorg Biochem, Volume 127, p.232-237, (2013)