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Projeto de investigação
Understanding enzymatic CO2 reduction for enhanced hybrid catalysts
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Tracking W-Formate Dehydrogenase Structural Changes During Catalysis and Enzyme Reoxidation
Publication . Vilela-Alves, Guilherme; Manuel, Rita Rebelo; Oliveira, Ana Rita; Pereira, Inês Cardoso; Romão, Maria João; Mota, Cristiano; DQ - Departamento de Química; UCIBIO - Applied Molecular Biosciences Unit; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); MDPI - Multidisciplinary Digital Publishing Institute
Metal-dependent formate dehydrogenases (Fdh) catalyze the reversible conversion of CO2 to formate, with unrivalled efficiency and selectivity. However, the key catalytic aspects of these enzymes remain unknown, preventing us from fully benefiting from their capabilities in terms of biotechnological applications. Here, we report a time-resolved characterization by X-ray crystallography of the Desulfovibrio vulgaris Hildenborough SeCys/W-Fdh during formate oxidation. The results allowed us to model five different intermediate structures and to chronologically map the changes occurring during enzyme reduction. Formate molecules were assigned for the first time to populate the catalytic pocket of a Fdh. Finally, the redox reversibility of DvFdhAB in crystals was confirmed by reduction and reoxidation structural studies.
Spectroscopic and Structural Characterization of Reduced Desulfovibrio vulgaris Hildenborough W-FdhAB Reveals Stable Metal Coordination during Catalysis
Publication . Oliveira, Ana Rita; Mota, Cristiano; Klymanska, Kateryna; Biaso, Frederic; Romão, Maria Joao; Guigliarelli, Bruno; Pereira, Inês Cardoso; Instituto de Tecnologia Química e Biológica António Xavier (ITQB); UCIBIO - Applied Molecular Biosciences Unit; DQ - Departamento de Química; ACS - American Chemical Society
Metal-dependent formate dehydrogenases are important enzymes due to their activity of CO2reduction to formate. The tungsten-containing FdhAB formate dehydrogenase from Desulfovibrio vulgaris Hildenborough is a good example displaying high activity, simple composition, and a notable structural and catalytic robustness. Here, we report the first spectroscopic redox characterization of FdhAB metal centers by EPR. Titration with dithionite or formate leads to reduction of three [4Fe-4S]1+clusters, and full reduction requires Ti(III)-citrate. The redox potentials of the four [4Fe-4S]1+centers range between -250 and -530 mV. Two distinct WVsignals were detected, WDVand WFV, which differ in only the g2-value. This difference can be explained by small variations in the twist angle of the two pyranopterins, as determined through DFT calculations of model compounds. The redox potential of WVI/Vwas determined to be -370 mV when reduced by dithionite and -340 mV when reduced by formate. The crystal structure of dithionite-reduced FdhAB was determined at high resolution (1.5 Å), revealing the same structural alterations as reported for the formate-reduced structure. These results corroborate a stable six-ligand W coordination in the catalytic intermediate WVstate of FdhAB.
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Entidade financiadora
Fundação para a Ciência e a Tecnologia
Programa de financiamento
3599-PPCDT
Número da atribuição
PTDC/BII-BBF/2050/2020