Autor(es): Campelo, Luísa Maria Ramos
Data: 2015
Identificador Persistente: http://hdl.handle.net/10362/15842
Origem: Repositório Institucional da UNL
Projeto/bolsa: info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FBBB-BEP%2F0753%2F2012/PT;
Assunto(s): Geobacter metallireducens; c -type cytochrome; Electron transfer; Protein cloning; Heterologous expression; Protein purification; Domínio/Área Científica::Engenharia e Tecnologia::Engenharia Química
Os resultados apresentados no capítulo 2 foram incluídos no artigo Dantas JM, Campelo LM, Duke NEC, Salgueiro CA, Pokkuluri PR (2015) "The structure of PccH from Geobacter sulfurreducens – a novel low reduction potential monoheme cytochrome essential for accepting electrons from an electrode", FEBS Journal, 282, 2215-2231.
The microorganims from the Geobacter genus are very interesting because of their potential use for biotechnological applications. They can transfer electrons towards extracellular terminal acceptors, which include toxic or radioactive metals. In addition, they also have the capability to convert renewable biomass into electricity. The most studied members of the Geobacter genus are the Geobacter metallireducens and Geobacter sulfurreducens bacteria. G. metallireducens displays some particular physiological aspects that are even more interesting compared to G. sulfurreducens, which include (i) their ability to reduce aromatic compounds, (ii) more efficient Fe(III) reduction rates, and (iii) the ability to use nitrate as terminal electron acceptor. Similarly, to G. sulfurreducens, a family of five homologous periplasmic triheme c-type cytochromes, designated PpcA, PpcB, PpcC, PpcE and PpcF, was identified in G. metallireducens. In G. sulfurreducens the triheme cytochromes were shown to be crucial for extracellular electron transfer and therefore it is conceivable that these proteins play a similar role in G. metallireducens. Therefore, the main goal of this work was to optimize the production of G. metallireducens triheme cytochromes as a foundation to future optimization of G. metallirudecens-based biotechnological applications. To achieve this goal, the proteins were cloned (except PpcB), heterologously expressed in Escherichia coli and purified. The preliminary characterization of PpcA and PpcF was also undertaken in the present work. The data obtained showed that both are low-spin hexacoordinated c-type cytochromes in the reduced and oxidized forms. In parallel, it was also undertaken an evolutionary study on PccH, a monoheme c-type cytochrome from G. sulfurreducens that is crucial for microbial electrosynthesis pathways in this bacterium. The results obtained suggested that PccH is a member of a new subclass within the class I cytochromes.
