STRUCTURAL AND MECHANISTIC FEATURES OF CYTOCHROME CD1 NITRITE REDUCTASE

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Development of new analytical tools for monitoring of cardiovascular disease markers – towards the detection of homocysteine-thiolactone

Publication . Monteiro, Tiago; Oliveira, Francisco; Silveira, Célia M.; Pereira, Sofia A.; Almeida, M. Gabriela

2015-05-10Conference object

Open access

Changes in metabolic pathways of Desulfovibrio alaskensis G20 cells induced by molybdate excess

Publication . Nair, Rashmi R.; Silveira, Célia M.; Diniz, Mário S.; Almeida, Maria G.; Moura, José J. G.; Rivas, Maria G.

The activity of sulfate-reducing bacteria (SRB) intensifies the problems associated to corrosion of metals and the solution entails significant economic costs. Although molybdate can be used to control the negative effects of these organisms, the mechanisms triggered in the cells exposed to Mo-excess are poorly understood. In this work, the effects of molybdate ions on the growth and morphology of the SRB Desulfovibrio alaskensis G20 (DaG20) were investigated. In addition, the cellular localization, ion uptake and regulation of protein expression were studied. We found that molybdate concentrations ranging between 50 and 150 μM produce a twofold increase in the doubling time with this effect being more significant at 200 μM molybdate (five times increase in the doubling time). It was also observed that 500 μM molybdate completely inhibits the cellular growth. On the context of protein regulation, we found that several enzymes involved in energy metabolism, cellular division and metal uptake processes were particularly influenced under the conditions tested. An overall description of some of the mechanisms involved in the DaG20 adaptation to molybdate-stress conditions is discussed.

2015-03Journal article

Restricted access

SERR spectroelectrochemical study of cytochrome cd1 nitrite reductase co-immobilized with physiological redox partner cytochrome c552 on biocompatible metal electrodes

Publication . Silveira, Célia M.; Quintas, Pedro O.; Moura, Isabel; Moura, José J. G.; Hildebrandt, Peter; Almeida, M. Gabriela; Todorovic, Smilja

"Cytochrome cd1 nitrite reductases (cd1NiRs) catalyze the one-electron reduction of nitrite to nitric oxide. Due to their catalytic reaction, cd1NiRs are regarded as promising components for biosensing, bioremediation and biotechnological applications. Motivated by earlier findings that catalytic activity of cd1NiR from Marinobacter hydrocarbonoclasticus (Mhcd1) depends on the presence of its physiological redox partner, cytochrome c552 (cyt c552), we show here a detailed surface enhanced resonance Raman characterization of Mhcd1 and cyt c552 attached to biocompatible electrodes in conditions which allow direct electron transfer between the conducting support and immobilized proteins. Mhcd1 and cyt c552 are co-immobilized on silver electrodes coated with self-assembled monolayers (SAMs) and the electrocatalytic activity of Ag // SAM // Mhcd1 // cyt c552 and Ag // SAM // cyt c552 // Mhcd1 constructs is tested in the presence of nitrite. Simultaneous evaluation of structural and thermodynamic properties of the immobilized proteins reveals that cyt c552 retains its native properties, while the redox potential of apparently intact Mhcd1 undergoes a ~150 mV negative shift upon adsorption. Neither of the immobilization strategies results in an active Mhcd1, reinforcing the idea that subtle and very specific interactions between Mhcd1 and cyt c552 govern efficient intermolecular electron transfer and catalytic activity of Mhcd1."

2015-06Journal article

Open access

New PON1-based biosensor for the detection of homocysteine-thiolactone in human plasma

Publication . Monteiro, Tiago; Oliveira, Francisco; Silveira, Célia M.; Pereira, Sofia A.; Almeida, M. Gabriela

2015-06-14Conference object

Open access

Small electron transfer proteins as mediators in enzymatic electrochemical biosensors

Publication . Silveira, Célia M.; Almeida, M. Gabriela

"Electrochemical mediators transfer redox equivalents between the active sites of enzymes and electrodes and, in this way, trigger bioelectrocatalytic redox processes. This has been very useful in the development of the so-called second generation biosensors, where they are able to transduce the catalytic event into an electrical signal. Among other pre-requisites, redox mediators must be readily oxidized/reduced at the electrode surface and easily interact with the biorecognition component. Small chemical compounds (e.g. ferrocene derivatives, ruthenium or osmium complexes and viologens) are frequently used for this purpose, but lately, small redox proteins (e.g. horse heart cytochrome c) have also played the role of redox partners in biosensing applications. In general, the docking between two complementary proteins introduces a second level of selectivity to the biosensor and enlarges the list of compounds targeted for analysis. Moreover, electrochemical interferences are frequently minimized owing to the small overpotentials achieved. This paper aims to provide an overview of enzyme biosensors that are mediated by electron transfer proteins. The article begins with a few considerations on mediated electrochemistry in biosensing 2 systems and proceeds with a detailed description of relevant works concerning the cooperative use of redox enzymes and biological electron donors/acceptors."

2013-04Journal article

Open access