Export 519 results:
Sort by: Author Title [ Type  (Asc)] Year
Journal Article
Topography of human cytochrome b5/cytochrome b5 reductase interacting domain and redox alterations upon complex formation, Samhan-Arias, A. K., Almeida R. M., Ramos S., Cordas C. M., Moura I., Gutierrez-Merino C., and Moura J. J. G. , Biochim Biophys Acta, Volume 1859, p.78-87, (2018)
Total lead and its stable isotopes in the digestive gland of Octopus vulgaris as a fingerprint, Raimundo, J., Vale C., Caetano M., Cesario R., and Moura I. , Aquatic Biology, 2009, Volume 6, Number 1-3, p.25-30, (2009) AbstractWebsite

We hypothesised that the isotopic signature of Pb in the digestive gland of the common octopus reflects the organisms' sources of Pb, and investigated whether isotopic Pb ratios are useful in characterising octopus populations. A total of 47 Octopus vulgaris individuals were captured between November 2005 and September 2006 in 2 areas of the Portuguese coast, near Matosinhos (Area A; NW coast) and Olhao (Area B; south coast), and digestive glands were analysed for total Pb and its stable isotopes. The same determinations were performed in 22 samples of surface sediments from the 2 areas. Pb concentrations in the digestive gland of specimens from Area B (2.8 to 13.0 mu g g(-1)) exceeded the values found in Area A (1.3 to 8.3 mu g g(-1)). A similar pattern was found for the isotopic Pb ratios: (206)Pb/(207)Pb was 1.173 to 1.185 for Area A and 1.165 to 1.172 for B; (206)Pb/(208)Pb was 0.476 to 0.487 for Area A and 0.318 to 0.483 for B. The different signatures of the digestive glands are in line with those observed in the surface sediments of the 2 coastal areas (e.g. (206)Pb/(207)Pb was 1.179 to 1.207 for Area A and 1.171 to 1.181 for B). However, the isotopic Pb signature of octopus was less radiogenic than that of sediments. Because octopus has a short life span (up to 24 mo) the signature reflects recent sources of Pb that have a less radiogenic signature. The Pb signature of surface sediments tends to integrate the record of the previous few years or decades, due to the frequent resuspension of the upper layer of coastal sediments. The mixing of sediments deposited during those periods results in higher isotopic Pb ratios (more radiogenic). The consistent differences between the 2 areas, in sediments and octopus, points towards the isotopic Pb signature as a possible useful tool to distinguish octopus populations.

Total synthesis of a simple metalloprotein-desulforedoxin, Tavares, P., Wunderlich J. K., Lloyd S. G., Legall J., Moura J. J., and Moura I. , Biochem Biophys Res Commun, Mar 17, Volume 208, Number 2, p.680-7, (1995) AbstractWebsite

Desulforedoxin is a protein purified from cellular extracts of Desulfovibrio gigas. It is a small (7.9 kDa) dimeric protein that contains a distorted rubredoxin like center (one single iron coordinated by four cysteinyl residues). Due to the simplicity of the polypeptide chain and of the iron center, an attempt was made to chemically produce this protein. A 36 amino acid polypeptide chain was synthesized based on the known sequence of native Desulforedoxin. The iron center was then reconstituted and the biochemical and spectroscopic characteristics of this synthetic protein were investigated. The final product has an equal sequence to the protein purified from D. gigas. The synthetic and natural Dx are very similar, in terms redox potential and spectroscopic properties (UV-Visible, EPR, Mossbauer).

Tungsten-containing formate dehydrogenase from Desulfovibrio gigas: metal identification and preliminary structural data by multi-wavelength crystallography, Raaijmakers, H., Teixeira S., Dias J. M., Almendra M. J., Brondino C. D., Moura I., Moura J. J., and Romao M. J. , J Biol Inorg Chem, Apr, Volume 6, Number 4, p.398-404, (2001) AbstractWebsite

The tungsten-containing formate dehydrogenase (W-FDH) isolated from Desulfovibrio gigas has been crystallized in space group P2(1), with cell parameters a = 73.8 A, b = 111.3 A, c = 156.6 A and beta = 93.7 degrees. These crystals diffract to beyond 2.0 A on a synchrotron radiation source. W-FDH is a heterodimer (92 kDa and 29 kDa subunits) and two W-FDH molecules are present in the asymmetric unit. Although a molecular replacement solution was found using the periplasmic nitrate reductase as a search model, additional phasing information was needed. A multiple-wavelength anomalous dispersion (MAD) dataset was collected at the W- and Fe-edges, at four different wavelengths. Anomalous and dispersive difference data allowed us to unambiguously identify the metal atoms bound to W-FDH as one W atom with a Se-cysteine ligand as well as one [4Fe-4S] cluster in the 92 kDa subunit, and three additional [4Fe-4S] centers in the smaller 29 kDa subunit. The D. gigas W-FDH was previously characterized based on metal analysis and spectroscopic data. One W atom was predicted to be bound to two molybdopterin guanine dinucleotide (MGD) pterin cofactors and two [4Fe-4S] centers were proposed to be present. The crystallographic data now reported reveal a selenium atom (as a Se-cysteine) coordinating to the W site, as well as two extra [4Fe-4S] clusters not anticipated before. The EPR data were re-evaluated in the light of these new results.

Two azurins with unusual redox and spectroscopic properties isolated from the Pseudomonas chlororaphis strains DSM 50083(T) and DSM 50135, Pinho, D., Besson S., Brondino C. D., Pereira E., de Castro B., and Moura I. , Journal of Inorganic Biochemistry, Feb, Volume 98, Number 2, p.276-286, (2004) AbstractWebsite

Two azurins (Az624 and Az626) were isolated from the soluble extract of two strains of Pseudomonas chlororaphis, DSM 50083(T) and DSM 50135, respectively, grown under microaerobic conditions with nitrate as final electron acceptor. The azurins, purified to electrophoretic homogeneity in three chromatographic steps, exhibit several peculiar properties. They have high reduction potentials and lower pI than most azurins described in the literature. As previously observed for Pseudomonas aeruginosa azurin, their reduction potentials are pH-dependent, but the pK values of their oxidized forms are lower, which suggests that deeper structural changes are associated with the oxidation process of these novel azurins. A hitherto undescribed pH-dependence of the diffusion coefficient was observed in Az624, that could be caused either by conformational changes, or by the formation of supramolecular aggregates associated with a protonation process. Both azurins exhibit axial X-band electron paramagnetic resonance spectra in frozen solution showing a typical hyperfine with the copper nucleus (I = 3/2) and a well-resolved superhyperfine structure with two equivalent N-14 nucleus (I = 1), which is not usually observed for azurins from other species. (C) 2003 Elsevier Inc. All rights reserved.

Two-dimensional 1H NMR studies on Desulfovibrio gigas ferredoxins. Assignment of the iron-sulfur cluster cysteinyl ligand protons, Macedo, Anjos L., Palma Nuno P., Moura Isabel, Legall Jean, Wray Victor, and Moura José J. G. , Magnetic Resonance in Chemistry, Volume 31, Number 13, p.S59-S67, (1993) AbstractWebsite

1D and 2D 1H NMR studies are reported on the oxidized and reduced [4Fe-4S] cluster of Desulfovibrio gigas ferredoxin I (Fdl). Several low-field contact shifted resonances (fast relaxing) are assigned to β-CH2 and α-CH coordinated cysteinyl residues. NOESY patterns (supported by 1D NOE experiments) resolves four pairs of geminal β-CH2 protons at low-field. The cluster ligands are assigned non-specifically to Cys8, Cys11, Cys14 and Cys50, based on the X-ray structural analysis available for the oligomeric form, FdII, that contains a single [3Fe-4S] cluster. It was indicated in this case that Cys11 is not bound to the trinuclear cluster but is tilted towards the solvent. The presence of four pairs of geminal β-CH2 protons for FdI unambiguously proves the occupancy of the fourth site of the [3Fe-4S] complex and implies the coordination of the Cys11 at the cluster. Analysis of the oxidized form of FdII, using the same methodology as described for FdI, supports the presence of three cysteinyl ligands in the [3Fe-4S] core. Further, the combined use of the X-ray coordinates enables the specific assignment of the three cysteinyl ligands of the cluster, extending a previous assignment of Cys50. In addition, very broad resonances were detected for the reduced form of FdII in the low-field region around 200 ppm and in the high field region around −80 ppm.

Ultrasonic assisted protein enzymatic digestion for fast protein identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry Sonoreactor versus ultrasonic probe, Rial-Otero, R., Carreira R. J., Cordeiro F. M., Moro A. J., Santos H. M., Vale G., Moura I., and Capelo J. L. , Journal of Chromatography A, Sep 28, Volume 1166, Number 1-2, p.101-107, (2007) AbstractWebsite

Two different ultrasonic energy sources, the sonoreactor and the ultrasonic probe, are compared for enzymatic digestion of proteins for protein identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDl-TOF-MS) using the peptide mass fingerprint (PMF) procedure. Variables such as (i) trypsin/protein ratio; (ii) sonication time; (iii) ultrasound amplitude; and (iv) protein concentration are studied and compared. As a general rule, the trypsin/protein ratio and the minimum protein concentration successfully digested are similar with both ultrasonic energy sources. Results showed that the time needed to digest proteins was shorter with the ultrasonic probe, 60 s versus 120 s, for the same amplitude of sonication, 50%. However, lower standard deviations and cleaner MALDI-TOF-MS spectra were obtained with the sonoreactor. In addition, the sonoreactor device provided higher sample throughput (6 samples for the sonoreactor versus 1 sample for the ultrasonic probe) and easier sample handling for lower sample volumes (25 mu l). Finally, a comparison of both methodologies for the specific identification of the adenylylsulphate reductase alfa subunit from a complex protein mixture from Desulfovibrio desulfuricans ATCC 27774 was done as a proof of the procedure. (c) 2007 Elsevier B.V. All rights reserved.

Ultrasonic multiprobe as a new tool to overcome the bottleneck of throughput in workflows for protein identification relaying on ultrasonic energy, Santos, H. M., Carreira R., Diniz M. S., Rivas M. G., Lodeiro C., Moura J. J., and Capelo J. L. , Talanta, Apr 15, Volume 81, Number 1-2, p.55-62, (2010) AbstractWebsite

We studied in this work the performance of the new ultrasonic multiprobe in terms of throughput, handling and robustness. The study was conducted using the multiprobe to speed two different proteomics workflows. The "classic" method relaying on overnight protein digestion (12h), was used as the standard procedure. This work clearly shows the importance of testing variables such as ultrasonic amplitude and ultrasonic time when adapting an ultrasonic-based treatment to a new ultrasonic device. The results here presented also shown and confirm the advantage of speed up sample treatment workflows with the aid of ultrasonic energy in combination with a 96-well plate. The methods compared were similar in terms of robustness, but the desalting free method was the fastest, requiring only 2 min/sample for completion. In addition it was also the simplest in terms of handling, since no desalting step was needed. The following standard proteins were successfully identified using the methods studied: bovine serum albumin, alpha-lactalbumin, ovalbumin, carbonic anhydrase, fructose-bisphosphate aldolase A, catalase, chymotrypsinogen A. As case study, the identification of the protein Split-Soret cytochrome c from D. desulfuricans ATCC 27774 was carried out.

Unambiguous identification of the nickel EPR signal in 61Ni-enriched Desulfovibrio gigas hydrogenase, Moura, J. J., Moura I., Huynh B. H., Kruger H. J., Teixeira M., DuVarney R. C., Dervartanian D. V., Xavier A. V., Peck, H. D. Jr., and Legall J. , Biochem Biophys Res Commun, Oct 29, Volume 108, Number 4, p.1388-93, (1982) AbstractWebsite
Understanding the response of Desulfovibrio desulfuricans ATCC 27774 to different electron acceptors - biosynthetic costs modulate substrate selection, Sousa, J. R., Silveira C. M., Fontes P., Roma-Rodrigues C., Fernandes A. R., Van Driessche G., Devreese B., Moura I., Moura J. J. G., and Almeida M. G. , Biochim Biophys Acta, Volume 1865, p.1455-1469, (2017)
Unusual reduction mechanism of copper in cysteine-rich environment, Maiti, B. K., Maia L., Moro A. J., Lima J. C., Cordas C., Moura I., and Moura J. J. G. , Inorg Chem, Volume 57, p.8078-8088, (2018) Website
Using cytochrome c(3) to make selenium nanowires, Abdelouas, A., Gong W. L., Lutze W., Shelnutt J. A., Franco R., and Moura I. , Chemistry of Materials, Jun, Volume 12, Number 6, p.1510-+, (2000) AbstractWebsite

We report on a new method to make nanostructures in aqueous solution at room temperature. We used the protein cytochrome c(3) to catalyze reduction of selenate (SeO42-) to selenium Se-0 by dithionite. Reduction was instantaneous. After a week spherical nanoparticles of red Se-0 (about 50 nm diameter) precipitated, followed by self-assembling into crystalline nanowires, typically 1 mu m long. The nanowires were composed of one strand of spherical particles; thicker strands contained several nanoparticles in parallel.

Vanadate oligomers interaction with phosphorylated myosin, Tiago, T., Aureliano M., Duarte R. O., and Moura J. J. G. , Inorganica Chimica Acta, Nov 15, Volume 339, p.317-321, (2002) AbstractWebsite

Using a myosin preparation containing endogenous myosin light-chain (LC2) kinase and phosphatase and calmodulin, i.e. near physiological ones, the interaction of vanadate oligomers with phosphorylated myosin was evaluated. Decavanadate or metavanadate solutions (2-15 mM total vanadate) did not prevent the phosphorylation state of the regulatory myosin lightchain, as observed by urea-polyacrylamide gel electrophoresis. The relative order of line broadening upon protein addition, reflecting the interaction of the vanadate oligomers with phosphorylated myosin, was V10 > V-4 > V-1 = 1 whereas, no changes were observed for monomeric vanadate. In the presence of ATP, V-1 signal was shifted upfield 2 ppm and became broadened, while V4 signal became narrowed. Moreover, a significant increase in myosin ATPase inhibition (60%) was observed when decameric vanadate species were present (1.4 mM). It is concluded that, under conditions near physiological ones, decameric vanadate differs from vanadate oligomers present in metavanadate solutions due to its strong interaction with the phosphorylated enzyme and myosin ATPase inhibition. Besides, ATP decreases the affinity of myosin for tetravanadate, induces the interaction with monomeric vanadate, whereas it does not affect decameric vanadate interaction. (C) 2002 Elsevier Science B.V. All rights reserved.

Vanadium distribution, lipid peroxidation and oxidative stress markers upon decavanadate in vivo administration, Soares, S. S., Martins H., Duarte R. O., Moura J. J., Coucelo J., Gutierrez-Merino C., and Aureliano M. , J Inorg Biochem, Jan, Volume 101, Number 1, p.80-8, (2007) AbstractWebsite

The contribution of decameric vanadate species to vanadate toxic effects in cardiac muscle was studied following an intravenous administration of a decavanadate solution (1mM total vanadium) in Sparus aurata. Although decameric vanadate is unstable in the assay medium, it decomposes with a half-life time of 16 allowing studying its effects not only in vitro but also in vivo. After 1, 6 and 12h upon decavanadate administration the increase of vanadium in blood plasma, red blood cells and in cardiac mitochondria and cytosol is not affected in comparison to the administration of a metavanadate solution containing labile oxovanadates. Cardiac tissue lipid peroxidation increases up to 20%, 1, 6 and 12h after metavanadate administration, whilst for decavanadate no effects were observed except 1h after treatment (+20%). Metavanadate administration clearly differs from decavanadate by enhancing, 12h after exposure, mitochondrial superoxide dismutase (SOD) activity (+115%) and not affecting catalase (CAT) activity whereas decavanadate increases SOD activity by 20% and decreases (-55%) mitochondrial CAT activity. At early times of exposure, 1 and 6h, the only effect observed upon decavanadate administration was the increase by 20% of SOD activity. In conclusion, decavanadate has a different response pattern of lipid peroxidation and oxidative stress markers, in spite of the same vanadium distribution in cardiac cells observed after decavanadate and metavanadate administration. It is suggested that once formed decameric vanadate species has a different reactivity than vanadate, thus, pointing out that the differential contribution of vanadium oligomers should be taken into account to rationalize in vivo vanadate toxicity.

A variable temperature spectroscopic study on Paracoccus pantotrophus pseudoazurin: Protein constraints on the blue Cu site, Xie, Xiangjin, Hadt Ryan G., Pauleta Sofia R., Gonzalez Pablo J., Un Sun, Moura Isabel, and Solomon Edward I. , Journal of Inorganic Biochemistry, Oct, Volume 103, Number 10, p.1307-1313, (2009) AbstractWebsite

The blue or Type 1 (T1) copper site of Paracoccus pantotrophus pseudoazurin exhibits significant absorption intensity in both the 450 and 600 nm regions. These are sigma and pi S(Cys) to Cu(2+) charge transfer (CT) transitions. The temperature dependent absorption, EPR, and resonance Raman (rR) vibrations enhanced by these bands indicate that a single species is present at all temperatures. This contrasts the temperature dependent behavior of the T1 center in nitrite reductase [S. Ghosh, X. Xie, A. Dey, Y. Sun, C. Scholes, E. Solomon, Proc. Natl. Acad. Sci. 106 (2009) 4969-4974] which has a thioether ligand that is unconstrained by the protein. The lack of temperature dependence in the T1 site in pseudoazurin indicates the presence of a protein constraint similar to the blue Cu site in plastocyanin where the thioether ligand is constrained at 2.8 angstrom. However, plastocyanin exhibits only pi CT. This spectral difference between pseudoazurin and plastocyanin reflects a coupled distortion of the site where the axial thiorether in pseudoazurin is also constrained, but at a shorter Cu-S(Met) bond length. This leads to an increase in the Cu(2+)-S(Cys) bond length, and the site undergoes a partial tetragonal distortion in pseudoazurin. Thus, its ground state wavefunction has both sigma and pi character in the Cu(2+)-S(Cys) bond. (C) 2009 Elsevier Inc. All rights reserved.

Voltammetric studies of the catalytic electron-transfer process between the Desulfovibrio gigas hydrogenase and small proteins isolated from the same genus, Moreno, C., Franco R., Moura I., Legall J., and Moura J. J. , Eur J Biochem, Nov 1, Volume 217, Number 3, p.981-9, (1993) AbstractWebsite

The kinetics of electron transfer between the Desulfovibrio gigas hydrogenase and several electron-transfer proteins from Desulfovibrio species were investigated by cyclic voltammetry, square-wave voltammetry and chronoamperometry. The cytochrome c3 from Desulfovibrio vulgaris (Hildenborough), Desulfovibrio desulfuricans (Norway 4), Desulfovibrio desulfuricans (American Type Culture Collection 27774) and D. gigas (NCIB 9332) were used as redox carriers. They differ in their redox potentials and isoelectric point. Depending on the pH, all the reduced forms of these cytochromes were effective in electron exchange with hydrogenase. Other small electron-transfer proteins such as ferredoxin I, ferredoxin II and rubredoxin from D. gigas were tentatively used as redox carriers. Only ferredoxin II was effective in mediating electron exchange between hydrogenase and the working electrode. The second-order rate constants k for the reaction between reduced proteins and hydrogenase were calculated based on the theory of the simplest electrocatalytic mechanism [Moreno, C., Costa, C., Moura, I., Le Gall, J., Liu, M. Y., Payne, W. J., van Dijk, C. & Moura, J. J. G. (1993) Eur. J. Biochem. 212, 79-86] and the results obtained by cyclic voltammetry were compared with those obtained by chronoamperometry. Values for k of 10(5)-10(6) M-1 s-1 (cytochrome c3 as electron carrier) and 10(4) M-1 s-1 (ferredoxin II as the electron carrier) were determined. The rate-constant values are discussed in terms of the existence of an electrostatic interaction between the electrode surface and the redox carrier and between the redox carrier and a positively charged part of the enzyme.

X-ray absorption spectroscopy of nickel in the hydrogenase from Desulfovibrio gigas, Scott, Robert A., Wallin Sten A., Czechowski Melvin, Dervartanian D. V., Legall Jean, Peck Harry D., and Moura Isabel , Journal of the American Chemical Society, 1984/10/01, Volume 106, Number 22, p.6864-6865, (1984) AbstractWebsite
X-ray crystal structure and EPR spectra of "arsenite-inhibited" Desulfovibriogigas aldehyde dehydrogenase: a member of the xanthine oxidase family, Boer, D. R., Thapper A., Brondino C. D., Romao M. J., and Moura J. J. , J Am Chem Soc, Jul 21, Volume 126, Number 28, p.8614-5, (2004) AbstractWebsite

X-ray crystallography has been used to determine the structure of arsenite-inhibited aldehyde dehydrogenase from Desulfovibrio gigas, a member of the xanthine oxidase family of mononuclear molybdenum enzymes. The structure shows an AsO3 moiety bound to the molybdenum atom of the active site through one of the oxygen atoms. A reduced sample of arsenite-inhibited aldehyde dehydrogenase has a Mo(V) signal that shows anisotropic hyperfine and quadrupole coupling to one arsenic atom. This signal has a strong resemblance with a previously reported signal for arsenite-inhibited xanthine oxidase.

Zinc-substituted Desulfovibrio gigas desulforedoxins: resolving subunit degeneracy with nonsymmetric pseudocontact shifts, Goodfellow, B. J., Nunes S. G., Rusnak F., Moura I., Ascenso C., Moura J. J., Volkman B. F., and Markley J. L. , Protein Sci, Oct, Volume 11, Number 10, p.2464-70, (2002) AbstractWebsite

Desulfovibrio gigas desulforedoxin (Dx) consists of two identical peptides, each containing one [Fe-4S] center per monomer. Variants with different iron and zinc metal compositions arise when desulforedoxin is produced recombinantly from Escherichia coli. The three forms of the protein, the two homodimers [Fe(III)/Fe(III)]Dx and [Zn(II)/Zn(II)]Dx, and the heterodimer [Fe(III)/Zn(II)]Dx, can be separated by ion exchange chromatography on the basis of their charge differences. Once separated, the desulforedoxins containing iron can be reduced with added dithionite. For NMR studies, different protein samples were prepared labeled with (15)N or (15)N + (13)C. Spectral assignments were determined for [Fe(II)/Fe(II)]Dx and [Fe(II)/Zn(II)]Dx from 3D (15)N TOCSY-HSQC and NOESY-HSQC data, and compared with those reported previously for [Zn(II)/Zn(II)]Dx. Assignments for the (13)C(alpha) shifts were obtained from an HNCA experiment. Comparison of (1)H-(15)N HSQC spectra of [Zn(II)/Zn(II)]Dx, [Fe(II)/Fe(II)]Dx and [Fe(II)/Zn(II)]Dx revealed that the pseudocontact shifts in [Fe(II)/Zn(II)]Dx can be decomposed into inter- and intramonomer components, which, when summed, accurately predict the observed pseudocontact shifts observed for [Fe(II)/Fe(II)]Dx. The degree of linearity observed in the pseudocontact shifts for residues >/=8.5 A from the metal center indicates that the replacement of Fe(II) by Zn(II) produces little or no change in the structure of Dx. The results suggest a general strategy for the analysis of NMR spectra of homo-oligomeric proteins in which a paramagnetic center introduced into a single subunit is used to break the magnetic symmetry and make it possible to obtain distance constraints (both pseudocontact and NOE) between subunits.