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Journal Article
Perturbation of membrane dynamics in nerve cells as an early event during bilirubin-induced apoptosis, Rodrigues, C. M., Sola S., Castro R. E., Laires P. A., Brites D., and Moura J. J. , J Lipid Res, Jun, Volume 43, Number 6, p.885-94, (2002) AbstractWebsite

Increased levels of unconjugated bilirubin, the end product of heme catabolism, impair crucial aspects of nerve cell function. In previous studies, we demonstrated that bilirubin toxicity may be due to cell death by apoptosis. To characterize the sequence of events leading to neurotoxicity, we exposed developing rat brain astrocytes and neurons to unconjugated bilirubin and investigated whether changes in membrane dynamic properties can mediate apoptosis. Bilirubin induced a rapid, dose-dependent increase in apoptosis, which was nevertheless preceded by impaired mitochondrial metabolism. Using spin labels and electron paramagnetic resonance spectroscopy analysis of whole cell and isolated mitochondrial membranes exposed to bilirubin, we detected major membrane perturbation. By physically interacting with cell membranes, bilirubin induced an almost immediate increase in lipid polarity sensed at a superficial level. The enhanced membrane permeability coincided with an increase in lipid fluidity and protein mobility and was associated with significant oxidative injury to membrane lipids. In conclusion, apoptosis of nerve cells induced by bilirubin is mediated by its primary effect at physically perturbing the cell membrane. Bilirubin directly interacts with membranes influencing lipid polarity and fluidity, protein order, and redox status. These data suggest that nerve cell membranes are primary targets of bilirubin toxicity.

The photochemical reaction between uranyl nitrate and azulene, Burrows, Hugh D., Cardoso Augusto C., Formosinho Sebastião J., Gil Ana M. P. C., da Miguel Maria Graça M., Barata Belamino, and J.G. Moura José , Journal of Photochemistry and Photobiology A: Chemistry, Volume 68, Number 3, p.279-287, (1992) AbstractWebsite
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The photochemical reaction between uranyl-nitrate and azulene, Burrows, H. D., Cardoso A. C., Formosinho S. J., Gil Ampc, Miguel M. D., Barata B., and Moura J. J. G. , Journal of Photochemistry and Photobiology a-Chemistry, Sep 30, Volume 68, Number 3, p.279-287, (1992) AbstractWebsite

On photolysis of solutions of azulene and uranyl nitrate in alcohols, a dark, amorphous precipitate is formed. Various analytical techniques show that this is a mixture of a uranium salt and an organic component, suggested to be polyazulene. The effects of various parameters on the yield of the product have been studied and it is found that oxygen facilitates the reaction. Electron spin resonance studies show that the product is paramagnetic, in agreement with the established ease of oxidation of polyazulene, and suggest that it is formed via electron transfer from azulene to excited uranyl ion, followed by successive dimerizations and deprotonations of radical cation intermediates.

Physico-chemical and Spectroscopic Properties of the Monohemic Cytochrome C552 from Pseudomonas nautica 617, Saraiva, Lígia M., Fauque Guy, Besson Stéphane, and Moura Isabel , European Journal of Biochemistry, Volume 224, Number 3, p.1011-1017, (1994) AbstractWebsite

A c-type monohemic ferricytochrome c552 (11 kDa) was isolated from the soluble extract of a marine denitrifier, Pseudomonas nautica strain 617, grown under anaerobic conditions with nitrate as final electron acceptor. The NH2-terminal sequence and the amino acid composition of the cytochrome were determined. The heme iron of the cytochrome c552 has histidine-methionine as axial ligands, and a pH-dependent mid-point redox potential, equal to 250 mV at pH 7.6. The presence of methionine was demonstrated by visible, EPR and NMR spectroscopies. The assignment of most of the hemic protons was performed applying two-dimensional NOE spectroscopy (NOESY), and the aromatic region was assigned through two-dimensional correlated spectroscopy (COSY) experiments. The EPR spectrum of the oxidised form of the cytochrome c552 is typical of a low-spin ferric heme.

Potential therapeutic approaches for a sleeping pathogen: tuberculosis a case for bioinorganic chemistry, Sousa, E. H. S., Diógenes I. C. N., Lopes L. G. F., and Moura J. J. G. , J Biol Inorg Chem, Volume 25, p.685, (2020)
Predicting Protein-Protein Interactions Using BiGGER: Case Studies, Almeida, R. M., Dell'Acqua S., Krippahl L., Moura J. J. G., and Pauleta S. R. , Molecules, Volume 21, p.1037, (2016) Website
Preliminary crystallographic analysis and further characterization of a dodecaheme cytochrome c from Desulfovibrio desulfuricans ATCC 27774, Coelho, A. V., Matias P. M., Sieker L. C., Morais J., Carrondo M. A., Lampreia J., Costa C., Moura J. J., Moura I., and Legall J. , Acta Crystallogr D Biol Crystallogr, Nov 1, Volume 52, Number Pt 6, p.1202-8, (1996) 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.

Preliminary crystallographic analysis of the oxidized form of a two mono-nuclear iron centres protein from Desulfovibrio desulfuricans ATCC 27774, Coelho, A. V., Matias P. M., Carrondo M. A., Tavares P., Moura J. J., Moura I., Fulop V., Hajdu J., and Legall J. , Protein Sci, Jun, Volume 5, Number 6, p.1189-91, (1996) 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.

The presence of redox-sensitive nickel in the periplasmic hydrogenase from Desulfovibrio gigas, Legall, J., Ljungdahl P. O., Moura I., Peck, H. D. Jr., Xavier A. V., Moura J. J., Teixera M., Huynh B. H., and Dervartanian D. V. , Biochem Biophys Res Commun, May 31, Volume 106, Number 2, p.610-6, (1982) AbstractWebsite
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Primary sequence, oxidation-reduction potentials and tertiary-structure prediction of Desulfovibrio desulfuricans ATCC 27774 flavodoxin, Caldeira, J., Palma P. N., Regalla M., Lampreia J., Calvete J., Schafer W., Legall J., Moura I., and Moura J. J. , Eur J Biochem, Mar 15, Volume 220, Number 3, p.987-95, (1994) AbstractWebsite

Flavodoxin was isolated and purified from Desulfovibrio desulfuricans ATCC 27774, a sulfate-reducing organism that can also utilize nitrate as an alternative electron acceptor. Mid-point oxidation-reduction potentials of this flavodoxin were determined by ultraviolet/visible and EPR methods coupled to potentiometric measurements and their pH dependence studied in detail. The redox potential E2, for the couple oxidized/semiquinone forms at pH 6.7 and 25 degrees C is -40 mV, while the value for the semiquinone/hydroquinone forms (E1), at the same pH, -387 mV. E2 varies linearly with pH, while E1 is independent of pH at high values. However, at low pH (< 7.0), this value is less negative, compatible with a redox-linked protonation of the flavodoxin hydroquinone. A comparative study is presented for Desulfovibrio salexigens NCIB 8403 flavodoxin [Moura, I., Moura, J.J.G., Bruschi, M. & LeGall, J. (1980) Biochim. Biophys. Acta 591, 1-8]. The complete primary amino acid sequence was obtained by automated Edman degradation from peptides obtained by chemical and enzymic procedures. The amino acid sequence was confirmed by FAB/MS. Using the previously determined tridimensional structure of Desulfovibrio vulgaris flavodoxin as a model [similarity, 48.6%; Watenpaugh, K.D., Sieker, L.C., Jensen, L.H., LeGall, J. & Dubourdieu M. (1972) Proc. Natl Acad. Sci. USA 69, 3185-3188], the tridimensional structure of D. desulfuricans ATCC 27774 flavodoxin was predicted using AMBER force-field calculations.

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

The primary structure of the beta subunit of Desulfovibrio desulfuricans (ATCC 27774) NiFe hydrogenase, Franco, R., Calvete J. J., Thole H. H., Raida M., Moura I., and Moura J. J. G. , Protein and Peptide Letters, Apr, Volume 4, Number 2, p.131-138, (1997) AbstractWebsite

The periplasmic [NiFe] hydrogenase isolated from Desulfovibrio (D.) desulfuricans (ATCC 27774) is a heterodimer of a 28 kDa (beta) and a 60 kDa (alpha) subunit. Here we report the complete amino acid sequence of the small (beta) polypeptide chain determined by Edman degradation of proteolytic fragments. Electrospray-ionization mass spectrometry of the native protein confirmed the sequencing results. The sequence is compared with that of D. gigas [NiFe] hydrogenase whose three-dimensional structure has been recently published.

The primary structure of the split-Soret cytochrome c from Desulfovibrio desulfuricans ATCC 27774 reveals an unusual type of diheme cytochrome c, Devreese, B., Costa C., Demol H., Papaefthymiou V., Moura I., Moura J. J., and Van Beeumen J. , Eur J Biochem, Sep 1, Volume 248, Number 2, p.445-51, (1997) AbstractWebsite

The complete amino acid sequence of the unusual diheme split-Soret cytochrome c from the sulphate-reducing Desulfovibrio desulfuricans strain ATCC 27774 has been determined using classical chemical sequencing techniques and mass spectrometry. The 247-residue sequence shows almost no similarity with any other known diheme cytochrome c, but the heme-binding site of the protein is similar to that of the cytochromes c3 from the sulphate reducers. The cytochrome-c-like domain of the protein covers only the C-terminal part of the molecule, and there is evidence for at least one more domain containing four cysteine residues, which might bind another cofactor, possibly a non-heme iron-containing cluster. This domain is similar to a sequence fragment of the genome of Archaeoglobus fulgidus, which confirms the high conservation of the genes involved in sulfate reduction.

Protein effects on the electronic structure of the [Fe4S4]2+ cluster in ferredoxin and HiPIP, Glaser, T., Bertini I., Moura J. J., Hedman B., Hodgson K. O., and Solomon E. I. , J Am Chem Soc, May 23, Volume 123, Number 20, p.4859-60, (2001) AbstractWebsite
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Proteins containing the factor F430 from methanosarcina barkeri and methanobacterium thermoautotrophicum: Isolation and properties, Moura, Isabel, Moura José J. G., Santos Helena, Xavier Antonio V., Burch Gary, Peck Jr Harry D., and Legall Jean , Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, Volume 742, Number 1, p.84-90, (1983) AbstractWebsite
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Proteins dominate in the surface layers formed on materials exposed to extracellular polymeric substances from bacterial cultures, Yang, Y., Wikieł A. J., Dall'agnol L. T., Eloy P., Genet M. J., Moura J. J. G., Sand W., Dupont-Gillain C. C., and Rouxhet P. G. , Biofouling, Volume 32, p.95-108, (2016)
Proteómica: a Interface entre a Biologia Molecular e a Biochemistry de Proteínas, Almeida, G., Rodrigues C., and Lampreia J. , Bol. Soc. Port. Química, Volume 82, p.49-56, (2001) Abstract
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Proton NMR spectra of rubredoxins: new resonances assignable to .alpha.-CH and .beta.-CH2 hydrogens of cysteinate ligands to iron(II), Werth, Mark T., Kurtz Donald M., Moura Isabel, and Legall Jean , Journal of the American Chemical Society, 1987/01/01, Volume 109, Number 1, p.273-275, (1987) AbstractWebsite
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Protonation state of the Cu4S2 CuZ site in nitrous oxide reductase: redox dependence and insight into reactivity, Johnston, E. M., Dell'Acqua S., Pauleta S. R., Moura I., and Solomon E. I. , Chem Sci, Volume 6, p.5670-5679, (2015)
Purification and characterization of a tungsten-containing formate dehydrogenase from Desulfovibrio gigas, Almendra, M. J., Brondino C. D., Gavel O., Pereira A. S., Tavares P., Bursakov S., Duarte R., Caldeira J., Moura J. J., and Moura I. , Biochemistry, Dec 7, Volume 38, Number 49, p.16366-72, (1999) AbstractWebsite

An air-stable formate dehydrogenase (FDH), an enzyme that catalyzes the oxidation of formate to carbon dioxide, was purified from the sulfate reducing organism Desulfovibrio gigas (D. gigas) NCIB 9332. D. gigas FDH is a heterodimeric protein [alpha (92 kDa) and beta (29 kDa) subunits] and contains 7 +/- 1 Fe/protein and 0.9 +/- 0.1 W/protein. Selenium was not detected. The UV/visible absorption spectrum of D. gigas FDH is typical of an iron-sulfur protein. Analysis of pterin nucleotides yielded a content of 1.3 +/- 0.1 guanine monophosphate/mol of enzyme, which suggests a tungsten coordination with two molybdopterin guanine dinucleotide cofactors. Both Mossbauer spectroscopy performed on D. gigas FDH grown in a medium enriched with (57)Fe and EPR studies performed in the native and fully reduced state of the protein confirmed the presence of two [4Fe-4S] clusters. Variable-temperature EPR studies showed the presence of two signals compatible with an atom in a d(1) configuration albeit with an unusual relaxation behavior as compared to the one generally observed for W(V) ions.

Purification and characterization of bisulfite reductase (desulfofuscidin) from Desulfovibrio thermophilus and its complexes with exogenous ligands, Fauque, G., Lino A. R., Czechowski M., Kang L., Dervartanian D. V., Moura J. J., Legall J., and Moura I. , Biochim Biophys Acta, Aug 1, Volume 1040, Number 1, p.112-8, (1990) AbstractWebsite

A dissimilatory bisulfite reductase has been purified from a thermophilic sulfate-reducing bacterium Desulfovibrio thermophilus (DSM 1276) and studied by EPR and optical spectroscopic techniques. The visible spectrum of the purified bisulfite reductase exhibits absorption maxima at 578.5, 392.5 and 281 nm with a weak band around 700 nm. Photoreduction of the native enzyme causes a decrease in absorption at 578.5 nm and a concomitant increase in absorption at 607 nm. When reduced, the enzyme reacts with cyanide, sulfite, sulfide and carbon monoxide to give stable complexes. The EPR spectrum of the native D. thermophilus bisulfite reductase shows the presence of a high-spin ferric signal with g values at 7.26, 4.78 and 1.92. Upon photoreduction the high-spin ferric heme signal disappeared and a typical 'g = 1.94' signal of [4Fe-4S] type cluster appeared. Chemical analyses show that the enzyme contains four sirohemes and eight [4Fe-4S] centers per mol of protein. The molecular mass determined by gel filtration was found to be 175 kDa. On SDS-gel electrophoresis the enzyme presents a main band of 44 to 48 kDa. These results suggest that the bisulfite reductase contains probably one siroheme and two [4Fe-4S] centers per monomer. The dissimilatory bisulfite reductase from D. thermophilus presents some homologous properties with desulfofuscidin, the bisulfite reductase isolated from Thermodesulfobacterium commune (Hatchikian, E.C. and Zeikus, J.G. (1983) J. Bacteriol. 153, 1211-1220).

Purification and characterization of desulfoferrodoxin. A novel protein from Desulfovibrio desulfuricans (ATCC 27774) and from Desulfovibrio vulgaris (strain Hildenborough) that contains a distorted rubredoxin center and a mononuclear ferrous center, Moura, I., Tavares P., Moura J. J., Ravi N., Huynh B. H., Liu M. Y., and Legall J. , J Biol Chem, Dec 15, Volume 265, Number 35, p.21596-602, (1990) AbstractWebsite

A new type of non-heme iron protein was purified to homogeneity from extracts of Desulfovibrio desulfuricans (ATCC 27774) and Desulfovibrio vulgaris (strain Hildenborough). This protein is a monomer of 16-kDa containing two iron atoms per molecule. The visible spectrum has maxima at 495, 368, and 279 nm and the EPR spectrum of the native form shows resonances at g = 7.7, 5.7, 4.1 and 1.8 characteristic of a high-spin ferric ion (S = 5/2) with E/D = 0.08. Mossbauer data indicates the presence of two types of iron: an FeS4 site very similar to that found in desulforedoxin from Desulfovibrio gigas and an octahedral coordinated high-spin ferrous site most probably with nitrogen/oxygen-containing ligands. Due to this rather unusual combination of active centers, this novel protein is named desulfoferrodoxin. Based on NH2-terminal amino acid sequence determined so far, the desulfoferrodoxin isolated from D. desulfuricans (ATCC 27774) appears to be a close analogue to a recently discovered gene product from D. vulgaris (Brumlik, M.J., and Voordouw, G. (1989) J. Bacteriol. 171, 49996-50004), which was suggested to be a rubredoxin oxidoreductase. However, reduced pyridine nucleotides failed to reduce the desulforedoxin-like center of this new protein.

Purification and characterization of three proteins from a halophilic sulfate-reducing bacterium,<i>Desulfovibrio salexigens</i&gt, Czechowski, M., Fauque G., Galliano N., Dimon B., Moura I., Moura J. J. G., Xavier A. V., Barato B. A. S., Lino A. R., and Legall J. , Journal of Industrial Microbiology & Biotechnology, Volume 1, Number 3, p.139-147, (1986) AbstractWebsite
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Purification and preliminary characterization of tetraheme cytochrome c3 and adenylylsulfate reductase from the peptidolytic sulfate-reducing bacterium Desulfovibrio aminophilus DSM 12254, Lopez-Cortes, A., Bursakov S., Figueiredo A., Thapper A. E., Todorovic S., Moura J. J., Ollivier B., Moura I., and Fauque G. , Bioinorg Chem Appl, p.81-91, (2005) AbstractWebsite

Two proteins were purified and preliminarily characterized from the soluble extract of cells (310 g, wet weight) of the aminolytic and peptidolytic sulfate-reducing bacterium Desulfovibrio (D.) aminophilus DSM 12254. The iron-sulfur flavoenzyme adenylylsulfate (adenosine 5'-phosphosulfate, APS) reductase, a key enzyme in the microbial dissimilatory sulfate reduction, has been purified in three chromatographic steps (DEAE-Biogel A, Source 15, and Superdex 200 columns). It contains two different subunits with molecular masses of 75 and 18 kDa. The fraction after the last purification step had a purity index (A(278nm) / A(388nm)) of 5.34, which was used for further EPR spectroscopic studies. The D. aminophilus APS reductase is very similar to the homologous enzymes isolated from D. gigas and D. desulfuricans ATCC 27774. A tetraheme cytochrome c(3) (His-heme iron-His) has been purified in three chromatographic steps (DEAE- Biogel A, Source 15, and Biogel-HTP columns) and preliminarily characterized. It has a purity index ([A(553nm) - A(570nm)](red) / A(280nm)) of 2.9 and a molecular mass of around 15 kDa, and its spectroscopic characterization (NMR and EPR) has been carried out. This hemoprotein presents similarities with the tetraheme cytochrome c(3) from Desulfomicrobium (Des.) norvegicum (NMR spectra, and N-terminal amino acid sequence).