Hydrogen production and deuterium-proton exchange reactions catalyzed by Desulfovibrio nickel(II)-substituted rubredoxins,
Saint-Martin, P., Lespinat P. A., Fauque G., Berlier Y., Legall J., Moura I., Teixeira M., Xavier A. V., and Moura J. J.
, Proc Natl Acad Sci U S A, Dec, Volume 85, Number 24, p.9378-80, (1988)
AbstractThe nickel tetrahedral sulfur-coordinated core formed upon metal replacement of the native iron in Desulfovibrio sp. rubredoxins is shown to mimic the reactivity pattern of nickel-containing hydrogenases with respect to hydrogen production, deuterium-proton exchange, and inhibition by carbon monoxide.
Peroxidase-like activity of cytochrome b5 is triggered upon hemichrome formation in alkaline pH,
Samhan-Arias, A., Maia L. B., Cordas C. M., Moura I., Gutierrez-Merino C., and Moura J. J. G.
, BBA - Proteins and Proteomics, Volume 1866, p.373-378, (2018)
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)
Ligand accessibility to heme cytochrome b5 coordinating sphere and enzymatic activity enhancement upon tyrosine ionization,
Samhan-Arias, A. K., Cordas C. M., Carepo M. S., Maia L. B., Gutierrez-Merino C., Moura I., and Moura J. J. G.
, J Biol Inorg Chem, Volume 24, p.317-330, (2019)
Cytochrome b5 reductase is the component from neuronal synaptic plasma membrane vesicles that generates superoxide anion upon stimulation by cytochrome c,
Samhan-Arias, A. K., Fortalezas S., Cordas C., Moura I., Moura J. J. G., and Gutierrez-Merino C.
, Redox Biol, Volume 15, p.109-114, (2018)
Reduction of ascorbate free radical by the plasma membrane of synaptic terminals from rat brain,
Samhan-Arias, A. K., Duarte R. O., Martin-Romero F. J., Moura J. J., and Gutierrez-Merino C.
, Arch Biochem Biophys, Jan 15, Volume 469, Number 2, p.243-54, (2008)
AbstractSynaptic plasma membranes (SPMV) decrease the steady state ascorbate free radical (AFR) concentration of 1mM ascorbate in phosphate/EDTA buffer (pH 7), due to AFR recycling by redox coupling between ascorbate and the ubiquinone content of these membranes. In the presence of NADH, but not NADPH, SPMV catalyse a rapid recycling of AFR which further lower the AFR concentration below 0.05 microM. These results correlate with the nearly 10-fold higher NADH oxidase over NADPH oxidase activity of SPMV. SPMV has NADH-dependent coenzyme Q reductase activity. In the presence of ascorbate the stimulation of the NADH oxidase activity of SPMV by coenzyme Q(1) and cytochrome c can be accounted for by the increase of the AFR concentration generated by the redox pairs ascorbate/coenzyme Q(1) and ascorbate/cytochrome c. The NADH:AFR reductase activity makes a major contribution to the NADH oxidase activity of SPMV and decreases the steady-state AFR concentration well below the micromolar concentration range.
Evidence for antisymmetric exchange in cuboidal 3Fe-4S (+) clusters,
Sanakis, Y., Macedo A. L., Moura I., Moura J. J. G., Papaefthymiou V., and Munck E.
, Journal of the American Chemical Society, Dec 6, Volume 122, Number 48, p.11855-11863, (2000)
AbstractIron-sulfur clusters with [3Fe-4S] cores are widely distributed in biological systems. In the oxidized state, designated [3Fe-4S](+), these electron-transfer agents have an electronic ground state with S = 1/2, and; they exhibit EPR signals centered at g = 2.01. It has been established by Mossbauer spectroscopy that the three iron sites of the cluster are high-spin Fe3+; and the general properties of the S = 1/2 ground state have been described with the exchange Hamiltonian H-exch = J(12)S(1).S-2 + J(23)S(2).S-3 + J(13)S(1).S-3 Some [3Fe-4S](+) clusters (type 1) have their g-values confined to the range between g = 2.03 and 2.00 while others (type 2) exhibit a continuous distribution of g-values down to g approximate to 1.85. Despite considerable efforts in various laboratories no model has emerged that explains the g-values of type 2 clusters. The 4.2 K spectra of all [3Fe-4S](+) clusters have broad features,which have been simulated in the past by using Fe-57 magnetic hyperfine tensors with anisotropies that are unusually large for high-spin feme sites. It is proposed here that antisymmetric exchange, H-AS = d.(S-1 x S-2 + S-2 x S-3 + S-3 x S-1), is the cause of the g-value shifts in type 2 clusters. We have been able to fit the EPR and Mossbauer spectra of the 3Fe clusters of beef heart aconitase and Desulfovibrio gigas ferredoxin II by using antisymmetric exchange in combination with distributed exchange coupling constants J(12), J(13), and J(23) (J-strain). While antisymmetric exchange is negligible for aconitase (which has a type 1 cluster), fits of the ferredoxin II spectra require \d\ approximate to 0.4 cm(-1). Our studies show that the data of both proteins can lie fit using the same isotropic Fe-57 magnetic hyperfine coupling constant for th three cluster sites, namely a -18.0 MHz for aconitase and a = -18.5 MHz for the D. gigas ferredoxin. The effects of antisymmetric exchange and J-strain on the Mossbauer and EPR spectra are discussed.
Electrochemical studies on small electron transfer proteins using membrane electrodes,
dos Santos, M. M. C., de Sousa P. M. P., Goncalves M. L. S., Krippahl L., Moura J. J. G., Lojou E., and Bianco P.
, Journal of Electroanalytical Chemistry, Jan 16, Volume 541, p.153-162, (2003)
AbstractMembrane electrodes (ME) were constructed using gold, glassy carbon and pyrolytic graphite supports and a dialysis membrane, and used to study the electrochemical behavior of small size electron transfer proteins: monohemic cytochrome c(522) from Pseudomonas nautica and cytochrome c(533) as well as rubredoxin from Desulfovibrio vulgaris. Different electrochemical techniques were used including cyclic voltammetry (CV), square wave voltammetry (SW) and differential pulse voltammetry (DP). A direct electrochemical response was obtained in all cases except with rubredoxin where a facilitator was added to the protein solution entrapped between the membrane and the electrode surface. Formal potentials and heterogeneous charge transfer rate constants were determined from the voltammetric data. The influence of the ionic strength and the pH of the medium on the electrochemical response at the ME were analyzed. The benefits from the use of the ME in protein electrochemistry and its role in modulating the redox behavior are analyzed. A critical comparison is presented with data obtained at non-MEs. Finally, the interactions that must be established between the proteins and the electrode surfaces are discussed, thereby modeling molecular interactions that occur in biological systems. (C) 2002 Elsevier Science B.V. All rights reserved.
Redox chemistry of low-pH forms of tetrahemic cytochrome c3,
Santos, M., Dos Santos M. M., Goncalves M. L., Costa C., Romao J. C., and Moura J. J.
, J Inorg Biochem, Dec, Volume 100, Number 12, p.2009-16, (2006)
AbstractDesulfovibrio vulgaris Hildenborough cytochrome c(3) contains four hemes in a low-spin state with bis-histidinyl coordination. High-spin forms of cytochrome c(3) can be generated by protonation of the axial ligands in order to probe spin equilibrium (low-spin/high-spin). The spin alterations occurring at acid pH, the associated changes in redox potentials, as well as the reactivity towards external ligands were followed by the conjunction of square wave voltammetry and UV-visible, CD, NMR and EPR spectroscopies. These processes may be used for modelling the action of enzymes that use spin equilibrium to promote enzyme activity and reactivity towards small molecules.
Synthesis of WO3 nanoparticles for biosensing applications,
Santos, L., Silveira C. M., Elangovan E., Neto J. P., Nunes D., Pereira L., Martins R., Viegas J., Moura J. J. G., Todorovic S., Almeida M. G., and Fortunato E. M.
, Sensors and Actuators B: Chemical, Volume 223, p.186-194, (2016)
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)
AbstractWe 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.
An improved clean sonoreactor-based method for protein identification by mass spectrometry-based techniques,
Santos, H. M., Mota Cristiano, Lodeiro C., Moura Isabel, Isaac Issa, and Capelo J. L.
, Talanta, Dec 15, Volume 77, Number 2, p.870-875, (2008)
AbstractA new clean fast (8 min) method for in-solution protein digestion Without detergent or urea for protein identification by peptide mass fingerprint and mass spectrometry-based techniques is Proposed. The new method avoids the use of time consuming desalting procedures entailing the following four steps done under the effect of an ultrasonic field provided by a sonoreactor: denaturation (1 min) in a mixed Solution of water:acetonitrile 1/1 (v/v): protein reduction (1 min); protein alkylation (1 min); and protein digestion (5 min). Five Proteins with masses comprised between 14.4 kDa and 97 kDa and the protein splitsoret cytochrome c from D. desulfuricans ATCC27774, Were Successfully identified with this procedure. No differences were found in the sequence coverage or in the number of peptides matched when the new clean method was compared to another one using urea. Twofold better signal-to-noise ratios were obtained in the MALDI spectra from protein samples prepared with the new method when comparing it with a method using urea. The new digestion method avoids the need to remove salt content and increases throughput (six samples at once) while reducing sample loss and contamination from sample handling. (C) 2008 Elsevier B.V. All rights reserved.