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Iron-Sulfur centers: Functions of an ancient metal site, Pauleta, S. R., Carepo M., Grazina R., Moura I., and Moura J. J. G. , Comprehensive Inorganic Chemistry III From Biology to Nanotechnology, vol. 2, ???, p.???, (2022)
Iron-sulfur centers: New roles for ancient metal sites, Grazina, R., Pauleta S., Moura J. J. G., and Moura I. , Comprehensive Inorganic Chemistry II, Vol. 3: Bioinorganic Fundamentals and Applications: Metals in Natural Living Systems and Metals in Toxicology and Medicine, Oxford, p.103-148, (2013)
Iron-sulphur cluster composition and redox properties of two ferredoxins from Desulfovibrio desulfuricans Norway strain, Guerlesquin, F., Moura J. J., and Cammack R. , Biochim Biophys Acta, Mar 16, Volume 679, Number 3, p.422-7, (1982) AbstractWebsite

Two ferredoxins from Desulfovibrio desulfuricans, Norway Strain, were investigated by EPR spectroscopy. Ferredoxin I appears to be a conventional [4Fe-4S]2+;1+ ferredoxin, with a midpoint reduction potential of -374 mV at pH 8. Ferredoxin II when reduced, at first showed a more complex spectrum, indicating an interaction between two [4Fe-4S] clusters, and probably, has two clusters per protein subunit. Upon reductive titration ferredoxin II changed to give a spectrum in which no intercluster interaction was seen. The midpoint potentials of the native and modified ferredoxin at pH 8 were estimated to be -500 and -440 mV, respectively.

Isolation and characterisation of a novel sulphate-reducing bacterium of the Desulfovibrio genus, Feio, M. J., Beech I. B., Carepo M., Lopes J. M., Cheung C. W., Franco R., Guezennec J., Smith J. R., Mitchell J. I., Moura J. J., and Lino A. R. , Anaerobe, Apr, Volume 4, Number 2, p.117-30, (1998) AbstractWebsite

A novel sulphate-reducing bacterium (Ind 1) was isolated from a biofilm removed from a severely corroded carbon steel structure in a marine environment. Light microscopy observations revealed that cells were Gram-negative, rod shaped and very motile. Partial 16S rRNA gene sequencing and analysis of the fatty acid profile demonstrated a strong similarity between the new species and members from the Desulfovibrio genus. This was confirmed by the results obtained following purification and characterisation of the key proteins involved in the sulphate-reduction pathway. Several metal-containing proteins, such as two periplasmic proteins: hydrogenase and cytochrome c3, and two cytoplasmic proteins: ferredoxin and sulphite reductase, were isolated and purified. The latter proved to be of the desulfoviridin type which is typical of the Desulfovibrio genus. The study of the remaining proteins revealed a high degree of similarity with the homologous proteins isolated from Desulfovibrio gigas. However, the position of the strain within the phylogenetic tree clearly indicates that the bacterium is closely related to Desulfovibrio gabonensis, and these three strains form a separate cluster in the delta subdivision of the Proteobacteria. On the basis of the results obtained, it is suggested that Ind 1 belongs to a new species of the genus Desulfovibrio, and the name Desulfovibrio indonensis is proposed.

Isolation and characterisation of metallothionein from the clam Ruditapes decussatus, Simes, D. C., Bebianno M. J., and Moura J. J. , Aquat Toxicol, May 8, Volume 63, Number 3, p.307-18, (2003) AbstractWebsite

Metallothioneins (MT) were obtained after purification from metal-exposed clams (Ruditapes decussatus) using gel-permeation and ion-exchange chromatography. Four cadmium-metallothioneins (CdMTs) were resolved by ion-exchange chromatography and they all had similar molecular weights, high cadmium content and an absorption spectra indicative of the presence of characteristic Cd-S aggregates. The NH(2)-terminal sequence suggests the presence of at least two class I clam MT isoforms. For the other two putative clam CdMTs isolated, the results of the amino acid determination were inconclusive. One was slightly contaminated and the other one had a blocked NH(2)-terminal. These clam metalothioneins contain glycine, which seems to be a common feature of molluscan MT family and exhibited more similarity to oysters than to mussels. Further investigation on the inducibility of these isoforms will be necessary if clams are to be used as biomarkers of metal exposure.

Isolation and characterization of a new Cu-Fe protein from Desulfovibrio aminophilus DSM12254, Rivas, M. G., Mota C. S., Pauleta S. R., Carepo M. S., Folgosa F., Andrade S. L., Fauque G., Pereira A. S., Tavares P., Calvete J. J., Moura I., and Moura J. J. , J Inorg Biochem, Oct, Volume 103, Number 10, p.1314-22, (2009) AbstractWebsite

The isolation and characterization of a new metalloprotein containing Cu and Fe atoms is reported. The as-isolated Cu-Fe protein shows an UV-visible spectrum with absorption bands at 320 nm, 409 nm and 615 nm. Molecular mass of the native protein along with denaturating electrophoresis and mass spectrometry data show that this protein is a multimer consisting of 14+/-1 subunits of 15254.3+/-7.6 Da. Mossbauer spectroscopy data of the as-isolated Cu-Fe protein is consistent with the presence of [2Fe-2S](2+) centers. Data interpretation of the dithionite reduced protein suggest that the metallic cluster could be constituted by two ferromagnetically coupled [2Fe-2S](+) spin delocalized pairs. The biochemical properties of the Cu-Fe protein are similar to the recently reported molybdenum resistance associated protein from Desulfovibrio, D. alaskensis. Furthermore, a BLAST search from the DNA deduced amino acid sequence shows that the Cu-Fe protein has homology with proteins annotated as zinc resistance associated proteins from Desulfovibrio, D. alaskensis, D. vulgaris Hildenborough, D. piger ATCC 29098. These facts suggest a possible role of the Cu-Fe protein in metal tolerance.

Isolation and characterization of a rubredoxin and a flavodoxin from Desulfovibrio desulfuricans Berre-Eau, Fauque, Guy D., Moura Isabel, Moura José J. G., Xavier António V., Galliano Nicole, and Legall Jean , Febs Letters, Volume 215, Number 1, p.63-67, (1987) AbstractWebsite
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Isolation and characterization of a rubredoxin and an (8Fe-8S) ferredoxin from Desulfuromonas acetoxidans, Probst, I., Moura J. J., Moura I., Bruschi M., and Legall J. , Biochim Biophys Acta, Apr 11, Volume 502, Number 1, p.38-44, (1978) AbstractWebsite

A two cluster (4Fe-4S) ferredoxin and a rubredoxin have been isolated from the sulfur-reducing bacterium Desulfuromonas acetoxidans. Their amino acid compositions are reported and compared to those of other iron-sulfur proteins. The ferredoxin contains 8 cysteine residues, 8 atoms of iron and 8 atoms of labile sulfur per molecule; its minimum molecular weight is 6163. The protein exhibits an abosrbance ratio of A385/A283 = 0.74. Storage results in a bleaching of the chromophore; the denatured ferredoxin is reconstitutable with iron and sulfide. The instability temperature is 52 degrees C. The rubredoxin does not differ markedly from rubredoxins from other anaerobic bacteria.

The isolation and characterization of cytochrome c nitrite reductase subunits (NrfA and NrfH) from Desulfovibrio desulfuricans ATCC 27774. Re-evaluation of the spectroscopic data and redox properties, Almeida, M. G., Macieira S., Goncalves L. L., Huber R., Cunha C. A., Romao M. J., Costa C., Lampreia J., Moura J. J., and Moura I. , Eur J Biochem, Oct, Volume 270, Number 19, p.3904-15, (2003) AbstractWebsite

The cytochrome c nitrite reductase is isolated from the membranes of the sulfate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 as a heterooligomeric complex composed by two subunits (61 kDa and 19 kDa) containing c-type hemes, encoded by the genes nrfA and nrfH, respectively. The extracted complex has in average a 2NrfA:1NrfH composition. The separation of ccNiR subunits from one another is accomplished by gel filtration chromatography in the presence of SDS. The amino-acid sequence and biochemical subunits characterization show that NrfA contains five hemes and NrfH four hemes. These considerations enabled the revision of a vast amount of existing spectroscopic data on the NrfHA complex that was not originally well interpreted due to the lack of knowledge on the heme content and the oligomeric enzyme status. Based on EPR and Mossbauer parameters and their correlation to structural information recently obtained from X-ray crystallography on the NrfA structure [Cunha, C.A., Macieira, S., Dias, J.M., Almeida, M.G., Goncalves, L.M.L., Costa, C., Lampreia, J., Huber, R., Moura, J.J.G., Moura, I. & Romao, M. (2003) J. Biol. Chem. 278, 17455-17465], we propose the full assignment of midpoint reduction potentials values to the individual hemes. NrfA contains the high-spin catalytic site (-80 mV) as well as a quite unusual high reduction potential (+150 mV)/low-spin bis-His coordinated heme, considered to be the site where electrons enter. In addition, the reassessment of the spectroscopic data allowed the first partial spectroscopic characterization of the NrfH subunit. The four NrfH hemes are all in a low-spin state (S = 1/2). One of them has a gmax at 3.55, characteristic of bis-histidinyl iron ligands in a noncoplanar arrangement, and has a positive reduction potential.

Isolation and characterization of desulforedoxin, a new type of non-heme iron protein from Desulfovibrio gigas, Moura, I., Bruschi M., Legall J., Moura J. J., and Xavier A. V. , Biochem Biophys Res Commun, Apr 25, Volume 75, Number 4, p.1037-44, (1977) AbstractWebsite
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Isolation and characterization of rubrerythrin, a non-heme iron protein from Desulfovibrio vulgaris that contains rubredoxin centers and a hemerythrin-like binuclear iron cluster, Legall, J., Prickril B. C., Moura I., Xavier A. V., Moura J. J., and Huynh B. H. , Biochemistry, Mar 8, Volume 27, Number 5, p.1636-42, (1988) AbstractWebsite

A new non-heme iron protein from the periplasmic fraction of Desulfovibrio vulgaris (Hildenbourough NCIB 8303) has been purified to homogeneity, and its amino acid composition, molecular weight, redox potential, iron content, and optical, EPR, and Mossbauer spectroscopic properties have been determined. This new protein is composed of two identical subunits with subunit molecular weight of 21,900 and contains four iron atoms per molecule. The as-purified oxidized protein exhibits an optical spectrum with absorption maxima at 492, 365, and 280 nm, and its EPR spectrum shows resonances at g = 4.3 and 9.4, characteristic of oxidized rubredoxin. The Mossbauer data indicate the presence of approximately equal amounts of two types of iron; we named them the Rd-like and the Hr-like iron due to their similarity to the iron centers of rubredoxins (Rds) and hemerythrins (Hrs), respectively. For the Rd-like iron, the measured fine and hyperfine parameters (D = 1.5 cm-1, E/D = 0.26, delta EQ = -0.55 mm/s, delta = 0.27 mm/s, Axx/gn beta n = -16.5 T, Ayy/gn beta n = -15.6 T, and Azz/gn beta n = -17.0 T) are almost identical with those obtained for the rubredoxin from Clostridium pasteurianum. Redox-titration studies monitored by EPR, however, showed that these Rd-like centers have a midpoint redox potential of +230 +/- 10 mV, approximately 250 mV more positive than those reported for rubredoxins. Another unusual feature of this protein is the presence of the Hr-like iron atoms.(ABSTRACT TRUNCATED AT 250 WORDS)

Isolation and preliminary characterization of a soluble nitrate reductase from the sulfate reducing organism Desulfovibrio desulfuricans ATCC 27774, Bursakov, S., Liu M. Y., Payne W. J., Legall J., Moura I., and Moura J. J. , Anaerobe, Feb, Volume 1, Number 1, p.55-60, (1995) AbstractWebsite

Desulfovibrio desulfuricans ATCC 27774 is a sulfate reducer that can adapt to nitrate respiration, inducing the enzymes required to utilize this alternative metabolic pathway. Nitrite reductase from this organism has been previously isolated and characterized, but no information was available on the enzyme involved in the reduction of nitrate. This is the first report of purification to homogeneity of a nitrate reductase from a sulfate reducing organism, thus completing the enzymatic system required to convert nitrate (through nitrite) to ammonia. D. desulfuricans nitrate reductase is a monomeric (circa 70 kDa) periplasmic enzyme with a specific activity of 5.4 K(m) for nitrate was estimated to be 20 microM. EPR signals due to one [4Fe-4S] cluster and Mo(V) were identified in dithionite reduced samples and in the presence of nitrate.

Isolation and spectroscopic characterization of the membrane-bound nitrate reductase from Pseudomonas chlororaphis DSM 50135, Pinho, D., Besson S., Silva P. J., de Castro B., and Moura I. , Biochimica Et Biophysica Acta-General Subjects, May 25, Volume 1723, Number 1-3, p.151-162, (2005) AbstractWebsite

A nitrate reductase was solubilized with Triton X-100 from the membranes of Pseudomonas chlororaphis DSM 50135 grown microaerobically in the presence of nitrate. Like other membrane-bound nitrate reductases, it contains three subunits, of 129, 66 (64) and 24 kDa, referred to in the literature as alpha, beta and gamma, respectively. Electrocatalytic studies revealed that only the membrane-bound, not the solubilized form of the enzyme, can accept electrons from a menaquinone analog, menadione, whereas both forms can accept electrons from methylviologen. The isolated enzyme possesses several iron-sulfur clusters and a molybdopterin guanine dinucleotide active center. The iron-sulfur clusters can be grouped in two classes according to their redox properties, the high-potential and low-potential clusters. In the as-isolated enzyme, two forms of the molybdenum center, high- and low-pH, are detectable by electron paramagnetic resonance spectroscopy. The low-pH form shows a hyperfine splitting due to a proton, suggesting the presence of an -OHx ligand. Dithionite reduces the Mo(V) center to Mo(W) and subsequent reoxidization with nitrate originates a new Mo(V) signal, identical to the oxidized low-pH form but lacking its characteristic hyperfine splitting. The isolated preparation also contains heme c (in a sub-stoichiometric amount) with the ability to relay electrons to the molybdenum center, suggesting that this nitrate reductase may contain heme c instead of the heme b usually found in this class of enzymes. (c) 2005 Elsevier B.V. All rights reserved.

Isolation of P590 from Methanosarcina barkeri: evidence for the presence of sulfite reductase activity, Moura, J. J., Moura I., Santos H., Xavier A. V., Scandellari M., and Legall J. , Biochem Biophys Res Commun, Oct 15, Volume 108, Number 3, p.1002-9, (1982) AbstractWebsite
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Isotropic exchange interaction between Mo and the proximal FeS center in the xanthine oxidase family member aldehyde oxidoreductase from Desulfovibrio gigas on native and polyalcohol inhibited samples: an EPR and QM/MM study, Gómez, M. C., Neuman N. I., Dalosto S. D., Gonzalez P. J., Moura J. J. G., Rizzi A. C., and Brondino C. D. , J Biol Inorg Chem, Volume 20, p.233–242, (2015)