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Fe-57 Q-band pulsed ENDOR of the hetero-dinuclear site of nickel hydrogenase: Comparison of the NiA, NiB, and NiC states, Huyett, J. E., Carepo M., Pamplona A., Franco R., Moura I., Moura J. J. G., and Hoffman B. M. , Journal of the American Chemical Society, Oct 1, Volume 119, Number 39, p.9291-9292, (1997) AbstractWebsite
Ferredoxin from Methanosarcina barkeri: evidence for the presence of a three-iron center, Moura, I., Moura J. J., Huynh B. H., Santos H., Legall J., and Xavier A. V. , Eur J Biochem, Aug, Volume 126, Number 1, p.95-8, (1982) AbstractWebsite

Methanosarcina barkeri ferredoxin was purified and characterized by electron paramagnetic resonance (EPR) and Mossbauer spectroscopy. The purification procedure included chromatographic steps on DEAE-cellulose and gel filtration. The isolated protein is unstable under aerobic conditions. The ferredoxin exhibits charge transfer bands at 283 nm and 405 nm with an absorption ratio A405/A283 = 0.73. Its molecular weight has been estimated to be 20000-22000 by gel filtration chromatography. The native ferredoxin exhibits an intense EPR signal at g = 2.02 and only a very weak g = 1.94 signal develops upon reduction with dithionite. The Mossbauer spectra of the reduced protein are characteristic of a [3Fe-3S] center. The combined EPR and Mossbauer studies show that M. barkeri ferredoxin contains only [3Fe-3S] clusters, similar to Azotobacter vinelandii Fd[Emptage, M.H., Kent, T.A., Huynh, B.H., Rawlings, J., Orme-Johnson, W.H. & Munck, M. (1980) J. Biol. Chem. 255, 1793-1796], Desulfovibrio gigas FdII [Huynh, B.H., Moura, J.J.G., Moura, I., Kent, T.A., LeGall, J., Xavier, A.V. & Munck, E. (1980) J. Biol. Chem. 255, 3242-3244] and mitochondrial beef heart aconitase [Kent, T.A., Dreyer, J.-L., Kennedy, M.C., Huynh, B.H., Emptage, M.H., Beinert, H. & Munck, E. (1982) Proc. Natl Acad. Sci. USA, 79, 1096-1100].

Ferredoxins, Moura, J. J., Macedo A. L., and Palma P. N. , Methods Enzymol, Volume 243, p.165-88, (1994) AbstractWebsite
Ferromagnetic resonance of Fe(111) thin films and Fe(111)/Cu(111) multilayers, Rezende, S. M., Moura J. A., de Aguiar F. M., and Schreiner W. H. , Phys Rev B Condens Matter, Jun 1, Volume 49, Number 21, p.15105-15109, (1994) AbstractWebsite
The first crystal structure of class III superoxide reductase from Treponema pallidum, Santos-Silva, T., Trincao J., Carvalho A. L., Bonifacio C., Auchere F., Raleiras P., Moura I., Moura J. J., and Romao M. J. , J Biol Inorg Chem, Jul, Volume 11, Number 5, p.548-58, (2006) AbstractWebsite

Superoxide reductase (SOR) is a metalloprotein containing a non-heme iron centre, responsible for the scavenging of superoxide radicals in the cell. The crystal structure of Treponema pallidum (Tp) SOR was determined using soft X-rays and synchrotron radiation. Crystals of the oxidized form were obtained using poly(ethylene glycol) and MgCl2 and diffracted beyond 1.55 A resolution. The overall architecture is very similar to that of other known SORs but TpSOR contains an N-terminal domain in which the desulforedoxin-type Fe centre, found in other SORs, is absent. This domain conserves the beta-barrel topology with an overall arrangement very similar to that of other SOR proteins where the centre is present. The absence of the iron ion and its ligands, however, causes a decrease in the cohesion of the domain and some disorder is observed, particularly in the region where the metal would be harboured. The C-terminal domain exhibits the characteristic immunoglobulin-like fold and harbours the Fe(His)4(Cys) active site. The five ligands of the iron centre are well conserved despite some disorder observed for one of the four molecules in the asymmetric unit. The participation of a glutamate as the sixth ligand of some of the iron centres in Pyrococcus furiosus SOR was not observed in TpSOR. A possible explanation is that either X-ray photoreduction occurred or there was a mixture of redox states at the start of data collection. In agreement with earlier proposals, details in the TpSOR structure also suggest that Lys49 might be involved in attraction of superoxide to the active site.

Flavodoxin and rubredoxin from Desulphovibrio salexigens, Moura, I., Moura J. J., Bruschi M., and Legall J. , Biochim Biophys Acta, Jun 10, Volume 591, Number 1, p.1-8, (1980) AbstractWebsite

A flavodoxin and a rubredoxin have been isolated from the sulfate-reducing bacterium Desulphovibrio salexigens (strain British Guiana, NICB 8403). Their amino acid composition and spectral characteristics did not differ markedly from the homologous proteins presented in other Desulphovibrio spp. Flavodoxin was shown to be active in the electron transport of the sulfite reductase system.

Fluorescence anisotropy of fluorescein varies according to pH: lessons for binding studies, Castro, N. S. S., Laia C. A. T., Moura I., and Carepo M. S. , J Photochem Photobiol A: Chemistry, Volume 372, p.59-62, (2019)
Formate dehydrogenase from Desulfovibrio desulfuricans ATCC 27774: Isolation and spectroscopic characterization of the active sites (heme, iron-sulfur centers and molybdenum), Costa, C., Teixeira M., Legall J., Moura J. J. G., and Moura I. , Journal of Biological Inorganic Chemistry, Apr, Volume 2, Number 2, p.198-208, (1997) AbstractWebsite

An air-stable formate dehydrogenase, an enzyme that catalyzes the oxidation of formate to CO2, was purified from a sulfate-reducing organism, Desulfovibrio desulfuricans ATCC 27774. The enzyme has a molecular mass of approximately 150 kDa (three different subunits: 88, 29 and 16 kDa) and contains three types of redox-active centers: four c-type hemes, nonheme iron arranged as two [4Fe-4S](2+/1+) centers and a molybdenum-pterin site. Selenium was also chemically detected. The enzyme specific activity is 78 units per mg of protein. Mo(V) EPR signals were observed in the native, reduced and formate-reacted states. EPR signals related to the presence of multiple low-spin hemes were also observed in the oxidized state. Upon reduction, an examination of the EPR data under appropriate conditions distinguishes two types of iron-sulfur centers, an [Fe-S] center I (g(max)=2.050, g(med)=1.947, g(min)=1.896) and an [Fe-S] center II (g(max)=2.071, g(med)=1.926, g(min)=1.865). Mossbauer spectroscopy confirmed the presence of four hemes in the low-spin state. The presence of two [4Fe-4S](2+/1+) centers was confirmed, one of these displaying very small hyperfine coupling constants in the +1 oxidation state. The midpoint redox potentials of the enzyme metal centers were also estimated.

The formate dehydrogenase isolated from the aerobe Methylobacterium sp. RXM is a molybdenum-containing protein, Duarte, R. O., Reis A. R., Girio F., Moura I., Moura J. J., and Collaco T. A. , Biochem Biophys Res Commun, Jan 3, Volume 230, Number 1, p.30-4, (1997) AbstractWebsite

The formate dehydrogenase (FDH) isolated from cells of Methylobacterium sp. RXM grown on molybdenum-containing mineral medium using methanol as carbon source, was partially purified (at least 90% pure as revealed by SDS-PAGE). The enzyme is unstable under oxygen and all the purification steps were conducted under strict anaerobic conditions. The molecular mass is 75 kDa (gel exclusion 300 kDa). The enzyme was characterized in terms of the kinetic parameters towards different substrates and electron acceptors, pH and temperature dependence and the effect of a wide range of compounds in the enzymatic activity. The EPR spectra of the dithionite reduced sample show, at low temperature (below 20 K), two rhombic EPR signals due to two distinct [Fe-S] centres (centre I at g-values 2.023, 1.951 and 1.933, and centre II at g-values 2.054 and 1.913). At high temperature (around 100 K) another rhombic EPR signal is optimally observed at g-values 2.002, 1.987 and 1.959 and attributed to the molybdenum site. The EPR signals assigned to the iron-sulfur centres show a strong analogy with the aldehyde oxido-reductase from Desulfovibrio gigas known to contain a Mo-pterin and two [2Fe-2S] centres and whose crystallographic structure was recently resolved.

Formation of a stable cyano-bridged dinuclear iron cluster following oxidation of the superoxide reductases from Treponema pallidum and Desulfovibrio vulgaris with K(3)Fe(CN)(6), Auchere, F., Raleiras P., Benson L., Venyaminov S. Y., Tavares P., Moura J. J., Moura I., and Rusnak F. , Inorg Chem, Feb 24, Volume 42, Number 4, p.938-40, (2003) AbstractWebsite

Superoxide reductases catalyze the monovalent reduction of superoxide anion to hydrogen peroxide. Spectroscopic evidence for the formation of a dinuclear cyano-bridged adduct after K(3)Fe(CN)(6) oxidation of the superoxide reductases neelaredoxin from Treponema pallidum and desulfoferrodoxin from Desulfovibrio vulgaris was reported. Oxidation with K(3)Fe(CN)(6) reveals a band in the near-IR with lambda(max) at 1020 nm, coupled with an increase of the iron content by almost 2-fold. Fourier transform infrared spectroscopy provided additional evidence with CN-stretching vibrations at 2095, 2025-2030, and 2047 cm(-)(1), assigned to a ferrocyanide adduct of the enzyme. Interestingly, the low-temperature electronic paramagnetic resonance (EPR) spectra of oxidized TpNlr reveal at least three different species indicating structural heterogeneity in the coordination environment of the active site Fe ion. Given the likely 6-coordinate geometry of the active site Fe(3+) ion in the ferrocyanide adduct, we propose that the rhombic EPR species can serve as a model of a hexacoordinate form of the active site.

Functional necessity and physicochemical characteristics of the 2Fe-2S cluster in mammalian ferrochelatase, Lloyd, S. G., Franco R., Moura J. J. G., Moura I., Ferreira G. C., and Huynh B. H. , Journal of the American Chemical Society, Oct 16, Volume 118, Number 41, p.9892-9900, (1996) AbstractWebsite

The recently discovered [2Fe-2S] cluster in mouse liver ferrochelatase has been characterized using UV-vis, EPR, and Mossbauer spectroscopic techniques. Studies are reported here for the recombinant protein purified from an overproducing transformed Escherichia coli strain. A positive correlation is observed between the presence of the [2Fe-2S] cluster and the enzymatic specific activity and demonstrates the necessity of this cofactor. Chemical analysis revealed that the preparations contained up to 1.3 Fe/molecule and indicated a 1:1 stoichiometry between Fe and acid-labile sulfide. The [2Fe-2S] cluster in the as-isolated ferrochelatase exhibits a UV-vis spectrum indicative of a [2Fe-2S](2+) cluster and is EPR-silent. The 8 T Mossbauer spectrum of the Fe-57-enriched as-isolated protein is well simulated by parameters Delta E(Q) = 0.69 +/- 0.03 mm/s and delta = 0.28 +/- 0.02 mm/s and confirms the presence of a diamagnetic ground state. Upon reduction with sodium dithionite, ferrochelatase shows a near-axial EPR spectrum with g-values of 2.00, 1.93, and 1.91, consistent with a S = 1/2 mixed valent Fe3+-Fe2+ cluster. The Orbach temperature dependence of the EPR line widths was used to provide an estimate of the exchange coupling J, which was determined to be on the order of 500-650 cm(-1) (+JS(1) . S-2 model). Redox titrations monitored by UV-vis and EPR spectroscopy revealed midpoint potentials of -390 +/- 10 and -405 +/- 10 mV, respectively. Mossbauer spectra of the sodium dithionite-reduced Fe-57-enriched ferrochelatase collected at 4.2 K in the presence of magnetic fields of 60 mT and 8 T strengths were analyzed in the mixed-valent S = 1/2 ground state. Parameters for the ferric site are Delta E(Q) = 1.2 +/- 0.2 mm/s and delta = 0.28 +/- 0.03 mm/s, with somewhat anisotropic hyperfine splittings; for the ferrous site, Delta E(Q) = 3.3 +/- 0.1 mm/s and delta = 0.67 +/- 0.04 mm/s with anisotropic hyperfine splittings characteristic of high-spin ferrous ion. The similarities and differences with other characterized [2Fe-2S](+) cluster-containing proteins are discussed.

The fundamental importance of basic science: examples of high-impact discoveries from an international Chemistry Network, Lopes, L. G. F., Sadler P. J., Bernardes-Génisson V., Moura J. J. G., Chauvin R., Bernhardt P. V., and Sousa E. H. S. , Quim Nova, Volume 43, p.1176-1189, (2020)
A further investigation of the cytochrome b5-cytochrome c complex, Banci, L., Bertini I., Felli I. C., Krippahl L., Kubicek K., Moura J. J., and Rosato A. , J Biol Inorg Chem, Sep, Volume 8, Number 7, p.777-86, (2003) AbstractWebsite

The interaction of reduced rabbit cytochrome b(5) with reduced yeast iso-1 cytochrome c has been studied through the analysis of (1)H-(15)N HSQC spectra, of (15)N longitudinal ( R(1)) and transverse ( R(2)) relaxation rates, and of the solvent exchange rates of protein backbone amides. For the first time, the adduct has been investigated also from the cytochrome c side. The analysis of the NMR data was integrated with docking calculations. The result is that cytochrome b(5) has two negative patches capable of interacting with a single positive surface area of cytochrome c. At low protein concentrations and in equimolar mixture, two different 1:1 adducts are formed. At high concentration and/or with excess cytochrome c, a 2:1 adduct is formed. All the species are in fast exchange on the scale of differences in chemical shift. By comparison with literature data, it appears that the structure of one 1:1 adduct changes with the origin or primary sequence of cytochrome b(5).

Gas chromatography mass spectrometry determination of acaricides from honey after a new fast ultrasonic-based solid phase micro-extraction sample treatment, Rial-Otero, R., Gaspar E. M., Moura I., and Capelo J. L. , Talanta, Mar 30, Volume 71, Number 5, p.1906-1914, (2007) AbstractWebsite

A method is reported for the determination of acaricides (amitraz, bromopropylate, coumaphos and fluvalinate) from honey by gas chromatography mass spectrometry after a new fast solid phase micro-extraction, SPME, procedure. Six different fibers were assessed for micro-extraction purpose studying the following variables: (i) SPME coating, (ii) extraction temperature, (iii) extraction time, (iv) desorption conditions and (v) agitation conditions. The new ultrasonic bath technology providing different sonication frequencies (35 and 130 kHz) and different working modes (Sweep, Standard and Degas) was studied and optimized for speeding up the acaricide micro-extraction. The best extraction results were achieved with the polyacrylate fiber. The extraction process was done in 30 min using the ultrasonic bath at 130 kHz in the Standard mode. Quality parameters of the proposed method show a good precision (<11%) and detection and quantitation limits lower than 6 and 15 ng/g, respectively, except for fluvalinate. Eleven Portuguese commercial honey samples were analyzed with the developed method in order to assess the performance of the method with real samples and to determine whether the concentration of acaricides in honey exceed their maximum residue levels (MRLs). Acaricide residues detected were lower than those established by the legislation. (c) 2006 Elsevier B.V. All rights reserved.

Gd(III) chelates as NMR probes of protein-protein interactions. Case study: rubredoxin and cytochrome c3, Almeida, R. M., Geraldes C. F., Pauleta S. R., and Moura J. J. , Inorg Chem, Nov 7, Volume 50, Number 21, p.10600-7, (2011) AbstractWebsite

Two cyclen-derived Gd probes, [Gd-DOTAM](3+) and [Gd-DOTP](5-) (DOTAM = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetamide; DOTP = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(methylenephosphonate)), were assessed as paramagnetic relaxation enhancement (PRE)-inducing probes for characterization of protein-protein interactions. Two proteins, Desulfovibrio gigas rubredoxin and Desulfovibrio gigas cytochrome c(3), were used as model partners. In a (1)H NMR titration it was shown that [Gd-DOTP](5-) binds to cytochrome c(3) near heme IV, causing pronounced PREs, characterized by line width broadenings of the heme methyl resonances at ratios as low as 0.08. A K(d) of 23 +/- 1 muM was calculated based on chemical shift perturbation of selected heme methyl resonances belonging to three different heme groups, caused by allosteric effects upon [Gd-DOTP](5-) binding to cytochrome c(3) at a molar ratio of 2. The other probe, [Gd-DOTAM](3+), caused PREs on a well-defined patch near the metal center of rubredoxin (especially the patch constituted by residues D19-G23 and W37-S45, which broaden beyond detection). This effect was partially reversed for some resonances (C6-Y11, in particular) when cytochrome c(3) was added to this system. Both probes were successful in causing reversible PREs at the partner binding site, thus showing to be good probes to identify partners' binding sites and since the interaction is reversible to structurally characterize protein complexes by better defining the complex interface.

Gene sequence and crystal structure of the aldehyde oxidoreductase from Desulfovibrio desulfuricans ATCC 27774, Rebelo, J., Macieira S., Dias J. M., Huber R., Ascenso C. S., Rusnak F., Moura J. J., Moura I., and Romao M. J. , J Mol Biol, Mar 17, Volume 297, Number 1, p.135-46, (2000) AbstractWebsite

The aldehyde oxidoreductase (MOD) isolated from the sulfate reducer Desulfovibrio desulfuricans (ATCC 27774) is a member of the xanthine oxidase family of molybdenum-containing enzymes. It has substrate specificity similar to that of the homologous enzyme from Desulfovibrio gigas (MOP) and the primary sequences from both enzymes show 68 % identity. The enzyme was crystallized in space group P6(1)22, with unit cell dimensions of a=b=156.4 A and c=177.1 A, and diffraction data were obtained to beyond 2.8 A. The crystal structure was solved by Patterson search techniques using the coordinates of the D. gigas enzyme. The overall fold of the D. desulfuricans enzyme is very similar to MOP and the few differences are mapped to exposed regions of the molecule. This is reflected in the electrostatic potential surfaces of both homologous enzymes, one exception being the surface potential in a region identifiable as the putative docking site of the physiological electron acceptor. Other essential features of the MOP structure, such as residues of the active-site cavity, are basically conserved in MOD. Two mutations are located in the pocket bearing a chain of catalytically relevant water molecules. As deduced from this work, both these enzymes are very closely related in terms of their sequences as well as 3D structures. The comparison allowed confirmation and establishment of features that are essential for their function; namely, conserved residues in the active-site, catalytically relevant water molecules and recognition of the physiological electron acceptor docking site.

Gene sequence and the 1.8 A crystal structure of the tungsten-containing formate dehydrogenase from Desulfovibrio gigas, Raaijmakers, H., Macieira S., Dias J. M., Teixeira S., Bursakov S., Huber R., Moura J. J., Moura I., and Romao M. J. , Structure, Sep, Volume 10, Number 9, p.1261-72, (2002) AbstractWebsite

Desulfovibrio gigas formate dehydrogenase is the first representative of a tungsten-containing enzyme from a mesophile that has been structurally characterized. It is a heterodimer of 110 and 24 kDa subunits. The large subunit, homologous to E. coli FDH-H and to D. desulfuricans nitrate reductase, harbors the W site and one [4Fe-4S] center. No small subunit ortholog containing three [4Fe-4S] clusters has been reported. The structural homology with E. coli FDH-H shows that the essential residues (SeCys158, His159, and Arg407) at the active site are conserved. The active site is accessible via a positively charged tunnel, while product release may be facilitated, for H(+) by buried waters and protonable amino acids and for CO(2) through a hydrophobic channel.

Genomic organization, gene expression and activity profile of Marinobacter hydrocarbonoclasticus denitrification enzymes, Carreira, C., Mestre O., Nunes R. F., Moura I., and Pauleta S. R. , PEERJ, Volume 6, p.DOI: 10.7717/peerj.5603, (2018)
Heterodimeric nitrate reductase (NapAB) from Cupriavidus necator H16: purification, crystallization and preliminary X-ray analysis, Coelho, C., Gonzalez P. J., Trincao J., Carvalho A. L., Najmudin S., Hettman T., Dieckman S., Moura J. J., Moura I., and Romao M. J. , Acta Crystallogr Sect F Struct Biol Cryst Commun, Jun 1, Volume 63, Number Pt 6, p.516-9, (2007) AbstractWebsite

The periplasmic nitrate reductase from Cupriavidus necator (also known as Ralstonia eutropha) is a heterodimer that is able to reduce nitrate to nitrite. It comprises a 91 kDa catalytic subunit (NapA) and a 17 kDa subunit (NapB) that is involved in electron transfer. The larger subunit contains a molybdenum active site with a bis-molybdopterin guanine dinucleotide cofactor as well as one [4Fe-4S] cluster, while the small subunit is a di-haem c-type cytochrome. Crystals of the oxidized form of this enzyme were obtained using polyethylene glycol 3350 as precipitant. A single crystal grown at the High Throughput Crystallization Laboratory of the EMBL in Grenoble diffracted to beyond 1.5 A at the ESRF (ID14-1), which is the highest resolution reported to date for a nitrate reductase. The unit-cell parameters are a = 142.2, b = 82.4, c = 96.8 A, beta = 100.7 degrees, space group C2, and one heterodimer is present per asymmetric unit.

Heteronuclear NMR and soft docking: an experimental approach for a structural model of the cytochrome c553-ferredoxin complex, Morelli, X., Dolla A., Czjzek M., Palma P. N., Blasco F., Krippahl L., Moura J. J., and Guerlesquin F. , Biochemistry, Mar 14, Volume 39, Number 10, p.2530-7, (2000) AbstractWebsite

The combination of docking algorithms with NMR data has been developed extensively for the studies of protein-ligand interactions. However, to extend this development for the studies of protein-protein interactions, the intermolecular NOE constraints, which are needed, are more difficult to access. In the present work, we describe a new approach that combines an ab initio docking calculation and the mapping of an interaction site using chemical shift variation analysis. The cytochrome c553-ferredoxin complex is used as a model of numerous electron-transfer complexes. The 15N-labeling of both molecules has been obtained, and the mapping of the interacting site on each partner, respectively, has been done using HSQC experiments. 1H and 15N chemical shift analysis defines the area of both molecules involved in the recognition interface. Models of the complex were generated by an ab initio docking software, the BiGGER program (bimolecular complex generation with global evaluation and ranking). This program generates a population of protein-protein docked geometries ranked by a scoring function, combining relevant stabilization parameters such as geometric complementarity surfaces, electrostatic interactions, desolvation energy, and pairwise affinities of amino acid side chains. We have implemented a new module that includes experimental input (here, NMR mapping of the interacting site) as a filter to select the accurate models. Final structures were energy minimized using the X-PLOR software and then analyzed. The best solution has an interface area (1037.4 A2) falling close to the range of generally observed recognition interfaces, with a distance of 10.0 A between the redox centers.

Hexaheme nitrite reductase from Desulfovibrio desulfuricans. Mossbauer and EPR characterization of the heme groups, Costa, C., Moura J. J., Moura I., Liu M. Y., Peck, H. D. Jr., Legall J., Wang Y. N., and Huynh B. H. , J Biol Chem, Aug 25, Volume 265, Number 24, p.14382-8, (1990) AbstractWebsite

Mossbauer and EPR spectroscopy were used to characterize the heme prosthetic groups of the nitrite reductase isolated from Desulfovibrio desulfuricans (ATCC 27774), which is a membrane-bound multiheme cytochrome capable of catalyzing the 6-electron reduction of nitrite to ammonia. At pH 7.6, the as-isolated enzyme exhibited a complex EPR spectrum consisting of a low-spin ferric heme signal at g = 2.96, 2.28, and 1.50 plus several broad resonances indicative of spin-spin interactions among the heme groups. EPR redox titration studies revealed yet another low-spin ferric heme signal at g = 3.2 and 2.14 (the third g value was undetected) and the presence of a high-spin ferric heme. Mossbauer measurements demonstrated further that this enzyme contained six distinct heme groups: one high-spin (S = 5/2) and five low-spin (S = 1/2) ferric hemes. Characteristic hyperfine parameters for all six hemes were obtained through a detailed analysis of the Mossbauer spectra. D. desulfuricans nitrite reductase can be reduced by chemical reductants, such as dithionite or reduced methyl viologen, or by hydrogenase under hydrogen atmosphere. Addition of nitrite to the fully reduced enzyme reoxidized all five low-spin hemes to their ferric states. The high-spin heme, however, was found to complex NO, suggesting that the high-spin heme could be the substrate binding site and that NO could be an intermediate present in an enzyme-bound form.

Highly sensitive nitrite biosensor based on the electrical wiring of nitrite reductase by ZnCr-AQS LDH, Chen, H., Mousty C., Cosnier S., Silveira C., Moura J. J. G., and Almeida M. G. , Electrochemistry Communications, Sep, Volume 9, Number 9, p.2240-2245, (2007) AbstractWebsite

A biosensor for amperometric determination of nitrite was developed using cytochrome c nitrite reductase (ccNiR) from Desulfovibrio desulfuricans immobilized and electrically connected on a glassy carbon electrode by entrapment into redox active [ZnCr-AQS] layered double hydroxide containing anthraquinone-2-sulfonate (AQS). The transduction step corresponded to the electro-enzymatic reduction of nitrite by immobilized AQS molecules at -0.6 V. The biosensor showed a fast response to nitrite (5 s) with a linear range between 0.015 and 2.35 mu M, a sensitivity of 1.8 A M-1 cm(-2) and a detection limit of 4 nM. The apparent Michaelis-Menten constant (K-M(app)) M was 7.5 mu M. (c) 2007 Elsevier B.V. All rights reserved.

How Biology handles nitrite, Maia, L., and Moura J. J. G. , Chem Rev, Volume 114, p.5273-5357, (2014)
Human erythrocytes exposure to juglone leads to an increase of superoxide anion production associated with cytochrome b5 reductase uncoupling, Valério, G. N., Gutierrez-Merino C., Nogueira F., Moura I., Moura J. J. G., and Samhan-Arias A. K. , Biochim Biophys Acta Bioenerg, Volume EPub, (2020)
Hydrogen evolution and consumption in AOT-isooctane reverse micelles by Desulfovibrio gigas hydrogenase, Andrade, S. L. A., and Moura J. J. G. , Enzyme and Microbial Technology, Sep 2, Volume 31, Number 4, p.398-402, (2002) AbstractWebsite

The enzyme hydrogenase isolated from the sulphate reducing anaerobic bacterium Desulfovibrio gigas was encapsulated in reverse micelles of AOT-water-isooctane. The enzyme ability to consume molecular hydrogen was studied as a function of the micelle size (given by W-o = [H2O]/[organic solvent]). A peak of catalytic activity was obtained for W-o = 18, a micelle size theoretically fitting the heterodimeric hydrogenase molecule. At this W-o value, the recorded catalytic activity was slightly higher than in a buffer system (K-cat = 169.43 s(-1) against the buffer value of 151 s(-1)). The optimal buffer used to encapsulate the enzyme was found to be imidazole 50 mM, pH 9.0, The molecular hydrogen production activity was also tested in this reverse micelle medium. (C) 2002 Elsevier Science Inc. All lights reserved.