ITQB: BEM - Artigos em revista internacional com arbitragem científica
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- Toward the Mechanistic Understanding of Enzymatic CO2 ReductionPublication . Oliveira, Ana Rita; Mota, Cristiano; Mourato, Cláudia; Domingos, Renato M.; Santos, Marino F. A.; Gesto, Diana; Guigliarelli, Bruno; Santos-Silva, Teresa; Romão, Maria João; Cardoso Pereira, Inês A.; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química; DCV - Departamento de Ciências da Vida; ACS - American Chemical SocietyReducing CO2 is a challenging chemical transformation that biology solves easily, with high efficiency and specificity. In particular, formate dehydrogenases are of great interest since they reduce CO2 to formate, a valuable chemical fuel and hydrogen storage compound. The metal-dependent formate dehydrogenases of prokaryotes can show high activity for CO2 reduction. Here, we report an expression system to produce recombinant W/Sec-FdhAB from Desulfovibrio vulgaris Hildenborough fully loaded with cofactors, its catalytic characterization and crystal structures in oxidized and reduced states. The enzyme has very high activity for CO2 reduction and displays remarkable oxygen stability. The crystal structure of the formate-reduced enzyme shows Sec still coordinating the tungsten, supporting a mechanism of stable metal coordination during catalysis. Comparison of the oxidized and reduced structures shows significant changes close to the active site. The DvFdhAB is an excellent model for studying catalytic CO2 reduction and probing the mechanism of this conversion.
- Proteomic and Isotopic Response of Desulfovibrio vulgaris to DsrC PerturbationPublication . Leavitt, William D.; Venceslau, Sofia S.; Waldbauer, Jacob; Smith, Derek A.; Cardoso Pereira, Inês A.; Bradley, Alexander S.; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); Frontiers Research FoundationDissimilatory sulfate reduction is a microbial energy metabolism that can produce sulfur isotopic fractionations over a large range in magnitude. Calibrating sulfur isotopic fractionation in laboratory experiments allows for better interpretations of sulfur isotopes in modern sediments and ancient sedimentary rocks. The proteins involved in sulfate reduction are expressed in response to environmental conditions, and are collectively responsible for the net isotopic fractionation between sulfate and sulfide. We examined the role of DsrC, a key component of the sulfate reduction pathway, by comparing wildtype Desulfovibrio vulgaris DSM 644T to strain IPFG07, a mutant deficient in DsrC production. Both strains were cultivated in parallel chemostat reactors at identical turnover times and cell specific sulfate reduction rates. Under these conditions, sulfur isotopic fractionations between sulfate and sulfide of 17.3 ± 0.5 or 12.6 ± 0.5 were recorded for the wildtype or mutant, respectively. The enzymatic machinery that produced these different fractionations was revealed by quantitative proteomics. Results are consistent with a cellular-level response that throttled the supply of electrons and sulfur supply through the sulfate reduction pathway more in the mutant relative to the wildtype, independent of rate. We conclude that the smaller fractionation observed in the mutant strain is a consequence of sulfate reduction that proceeded at a rate that consumed a greater proportion of the strains overall capacity for sulfate reduction. These observations have consequences for models of sulfate reducer metabolism and how it yields different isotopic fractionations, notably, the role of DsrC in central energy metabolism.
- Interfacing Formate Dehydrogenase with Metal Oxides for the Reversible Electrocatalysis and Solar-Driven Reduction of Carbon DioxidePublication . Miller, Melanie; Robinson, William E.; Oliveira, Ana Rita; Heidary, Nina; Kornienko, Nikolay; Warnan, Julien; Pereira, Inês A.C.; Reisner, Erwin; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); John Wiley & Sons, Ltd.The integration of enzymes with synthetic materials allows efficient electrocatalysis and production of solar fuels. Here, we couple formate dehydrogenase (FDH) from Desulfovibrio vulgaris Hildenborough (DvH) to metal oxides for catalytic CO 2 reduction and report an in-depth study of the resulting enzyme–material interface. Protein film voltammetry (PFV) demonstrates the stable binding of FDH on metal-oxide electrodes and reveals the reversible and selective reduction of CO 2 to formate. Quartz crystal microbalance (QCM) and attenuated total reflection infrared (ATR-IR) spectroscopy confirm a high binding affinity for FDH to the TiO 2 surface. Adsorption of FDH on dye-sensitized TiO 2 allows for visible-light-driven CO 2 reduction to formate in the absence of a soluble redox mediator with a turnover frequency (TOF) of 11±1 s −1 . The strong coupling of the enzyme to the semiconductor gives rise to a new benchmark in the selective photoreduction of aqueous CO 2 to formate.
- Photoreduction of CO2 with a Formate Dehydrogenase Driven by Photosystem II Using a Semi-artificial Z-Scheme ArchitecturePublication . Sokol, Katarzyna P.; Robinson, William E.; Oliveira, Ana Rita; Warnan, Julien; Nowaczyk, Marc M.; Ruff, Adrian; Pereira, Inês A.C.; Reisner, Erwin; Bioresources 4 Sustainability (GREEN-IT); Molecular, Structural and Cellular Microbiology (MOSTMICRO); Instituto de Tecnologia Química e Biológica António Xavier (ITQB); ACS - American Chemical SocietySolar-driven coupling of water oxidation with CO2 reduction sustains life on our planet and is of high priority in contemporary energy research. Here, we report a photoelectrochemical tandem device that performs photocatalytic reduction of CO2 to formate. We employ a semi-artificial design, which wires a W-dependent formate dehydrogenase (FDH) cathode to a photoanode containing the photosynthetic water oxidation enzyme, Photosystem II, via a synthetic dye with complementary light absorption. From a biological perspective, the system achieves a metabolically inaccessible pathway of light-driven CO2 fixation to formate. From a synthetic point of view, it represents a proof-of-principle system utilizing precious-metal-free catalysts for selective CO2-to-formate conversion using water as an electron donor. This hybrid platform demonstrates the translatability and versatility of coupling abiotic and biotic components to create challenging models for solar fuel and chemical synthesis.
- Reversible and Selective Interconversion of Hydrogen and Carbon Dioxide into Formate by a Semiartificial Formate Hydrogenlyase MimicPublication . Sokol, Katarzyna P.; Robinson, William E.; Oliveira, Ana R.; Zacarias, Sonia; Lee, Chong Yong; Madden, Christopher; Bassegoda, Arnau; Hirst, Judy; Pereira, Inês A.C.; Reisner, Erwin; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); ACS - American Chemical SocietyThe biological formate hydrogenlyase (FHL) complex links a formate dehydrogenase (FDH) to a hydrogenase (H2ase) and produces H2 and CO2 from formate via mixed-acid fermentation in Escherichia coli. Here, we describe an electrochemical and a colloidal semiartificial FHL system that consists of an FDH and a H2ase immobilized on conductive indium tin oxide (ITO) as an electron relay. These in vitro systems benefit from the efficient wiring of a highly active enzyme pair and allow for the reversible conversion of formate to H2 and CO2 under ambient temperature and pressure. The hybrid systems provide a template for the design of synthetic catalysts and surpass the FHL complex in vivo by storing and releasing H2 on demand by interconverting CO2/H2 and formate with minimal bias in either direction.
- A gas breathing hydrogen/air biofuel cell comprising a redox polymer/hydrogenase-based bioanodePublication . Szczesny, Julian; Marković, Nikola; Conzuelo, Felipe; Zacarias, Sónia; Pereira, Inês A.C.; Lubitz, Wolfgang; Plumeré, Nicolas; Schuhmann, Wolfgang; Ruff, Adrian; Bioresources 4 Sustainability (GREEN-IT); Molecular, Structural and Cellular Microbiology (MOSTMICRO); Instituto de Tecnologia Química e Biológica António Xavier (ITQB); Nature PortfolioHydrogen is one of the most promising alternatives for fossil fuels. However, the power output of hydrogen/oxygen fuel cells is often restricted by mass transport limitations of the substrate. Here, we present a dual-gas breathing H2/air biofuel cell that overcomes these limitations. The cell is equipped with a hydrogen-oxidizing redox polymer/hydrogenase gas-breathing bioanode and an oxygen-reducing bilirubin oxidase gas-breathing biocathode (operated in a direct electron transfer regime). The bioanode consists of a two layer system with a redox polymer-based adhesion layer and an active, redox polymer/hydrogenase top layer. The redox polymers protect the biocatalyst from high potentials and oxygen damage. The bioanodes show remarkable current densities of up to 8 mA cm-2. A maximum power density of 3.6 mW cm-2 at 0.7 V and an open circuit voltage of up to 1.13 V were achieved in biofuel cell tests, representing outstanding values for a device that is based on a redox polymer-based hydrogenase bioanode.
- An electrogenic redox loop in sulfate reductionreveals a likely widespread mechanism of energy conservationPublication . G. Duarte, Américo; Catarino, Teresa; White, Gaye F.; Lousa, Diana; Neukirchen, Sinje; Soares, Cláudio; Sousa, Filipa L; Clarke, Thomas A.; Pereira, Inês A. C.The bioenergetics of anaerobic metabolism frequently relies on redox loops performed by membrane complexes with substrate- and quinone-binding sites on opposite sides of the membrane. However, in sulfate respiration (a key process in the biogeochemical sulfur cycle), the substrate- and quinone-binding sites of the QrcABCD complex are periplasmic, and their role in energy conservation has not been elucidated. Here we show that the QrcABCD complex of Desulfovibrio vulgaris is electrogenic, as protons and electrons required for quinone reduction are extracted from opposite sides of the membrane, with a H+/e− ratio of 1. Although the complex does not act as a H+-pump, QrcD may include a conserved proton channel leading from the N-side to the P-side menaquinone pocket. Our work provides evidence of how energy is conserved during dissimilatory sulfate reduction, and suggests mechanisms behind the functions of related bacterial respiratory complexes in other bioenergetic contexts. (...)
- Product quality of a recombinant fusion protein expressed in immobilised baby hamster kidney cells grown in protein-free mediumPublication . Cruz, H. J.; Dias, E. M.; Peixoto, C. M.; Moreira, J. L.; Carrondo, Manuel J. T.