ITQB: MMB - MA Dissertations
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- Unravelling the catalytic mechanism for NO reduction by a flavodiiron protein from Escherichia coliPublication . Oliveira, Fabiana Raquel Silva; Teixeira, Miguel; Folgosa, Filipe"Flavodiiron proteins (FDPs) have been identified as participants in the response mechanisms to nitrosative and oxidative stresses in strict and facultative anaerobic microorganisms. These proteins are composed of a domain containing the differic catalytic center and another containing an FMN. Escherichia coli, a human pathogen, encodes for a class B FDP (named flavorubredoxin, FlRd), having an extra rubredoxin center. FlRd is the only FDP described to date as having NO as its main substrate. Thus, this work aimed to understand the catalytic mechanism of E. coli FDP reducing NO to generate N2O.(...)"
- Superoxide reductases: unravelling catalytic determinants through natural and site directed mutantsPublication . Mordido, V. H.; Folgosa, Filipe; Teixeira, Miguel"Superoxide reductases (SOR) are non-heme iron enzymes able to catalyze the one-electron reduction of superoxide (O •), producing hydrogen peroxide (HO), with the assistance of an electron donor (usually a rubredoxin). This family of enzymes is usually divided into two classes: 2Fe-SOR, if they have two iron sites – centre I, similar to that found in desulforedoxin with the iron being coordinated by four cysteines; and centre II, the catalytic site, where the iron is coordinated by four equatorial histidines and one axial cysteine; or 1Fe-SOR, containing only the catalytic centre II.(...)"
- Exploring respiratory enzymes from Staphylococcus aureusPublication . Barbosa, Catarina; Pereira, Manuela; Refojo, Ana"Staphylococcus aureus is an opportunistic pathogen that can cause various disease patterns due to its high adaptative capacity. Many aspects of the energy metabolism and its respiratory chain system are still poorly understood. Herein, we report our study of three respiratory enzymes: Membrane potential- generating system (MpsAB) and two malate:quinone oxidoreductases (MQO). The MpsAB enzyme is a membrane complex proposed to be involved in energy conservation in S. aureus due the sequence homology of the subunit MpsA with the proton-translocation subunit NuoL of complex I from Escherichia coli. To further investigate the complex a series of expression tests were made to achieve its heterologous expression in E. coli. We also tried to optimize protein purification for biochemical studies. (...)"
- Insights into determinants of the activity of the flavodiiron NO reductase from Escherichia coliPublication . Soares, Jéssica; Teixeira, Miguel"Flavodiiron proteins (FDPs) are a family of metalloproteins involved in the reduction of molecular oxygen and hydrogen peroxide to water and/or the reduction of nitric oxide to nitrous oxide. Bioinformatic analysis of the primary structure of the FDPs led to the identification of a conserved motif in class B FDPs (flavorubredoxins, FlRd), that is relatively close to the active centre. The present experimental work aimed to study the role played by two serines present in this motif in an FDP from E.coli. Thus, two mutants, S33D and S34D, were studied and characterized in order to determine the implications of such mutations on the biochemical, kinetic and spectroscopic properties of the enzyme. The production of the mutants was achieved in E. coli BL21 (DE3) Gold and it was found that the biochemical characteristics were kept almost unchanged.(...)"
- Molecular and Cellular Investigation of Malate:quinone oxidoreductases from Staphylococcus aureusPublication . Gonçalves, Joana Lisboa da Silva; Pereira, Manuela"Staphylococcus aureus are opportunistic pathogens and represent one of the most frequent causes for community acquired and nosocomial infections. Over time, S. aureus has developed several mechanisms which led to the selection of increasingly resistant strains. These potentially lethal bacterial pathogens have become a major public health threat, being urgent the development of new drugs against this worldwide problem. However, and intriguingly many fundamental aspects of S. aureus have escaped attention, such as its energy metabolism and respiratory enzymes. We aim to contribute to this knowledge by exploring malate:quinone oxidoreductases at molecular and cellular levels.(...)"