SCAN: Biochemical and stability studies of hyperthermophilic multicopper oxidases

SCAN Seminar

Title: Biochemical and stability studies of hyperthermophilic multicopper oxidases

Speaker: André Fernandes, Microbial & Enzyme Technology Lab

Abstract:

The multicopper oxidases (MCOs) constitute a family of enzymes that present broad substrate specificity, oxidizing numerous aromatic phenols and amines. This family of enzymes is widely distributed throughout nature. The one-electron oxidation of these substrates occurs concomitantly with a four-electron reduction of molecular oxygen to water. The reactions catalyzed by these enzymes depend on the presence of two copper centres; a mononuclear type 1 copper centre that is the primary acceptor for electrons; and a trinuclear centre comprising one type 2 and two type 3 copper ions that are involved in dioxygen reduction to water. The laccases constitute a large subfamily of MCOs and have a great potential in various biotechnological processes mainly owing to their high relative nonspecific oxidation capacity, the lack of a requirement for cofactors and the use of readily available oxygen as an electron acceptor. A few MCO members are able to oxidize lower valence metal ions, such as Cu+, Fe2+ and Mn2+, with high specificity, and thus designated as metallo-oxidases such as human ceruloplasmin, yeast ferroxidase Fet3p and CueO from Escherichia coli, that are known to be critically involved in metal homeostasis mechanisms. The catalytic and stability characteristics of bacterial laccases at the molecular level are of considerable interest and, as a model system, the CotA-laccase from Bacillus subtilis has been extensively studied by our group. One of the objectives of such studies is to dissect the stability determinants using protein engineering techniques and to design MCO’s that better match biotechnological applications. These stability studies were extended to hyperthermophilic MCO’s. We have cloned, expressed, purified and studied two multicopper oxidases from hyperthermophilic microorganisms, McoA from Aquifex aeolicus and McoP from the Pyrobaculum aerophilum (the first MCO from Archaea). Both proteins present higher activities for metal ions, as compared to aromatic compounds, and melting temperatures of around 100ºC. McoA biochemical, structural and genetic properties suggest a role in the organism metal homeostasis. McoP has a low redox potential among MCO’s and could be involved in the denitrification pathway due to fact that it uses nitrous oxide as electron acceptor as well as oxygen.

Short CV:

2000-2005: Degree in Biotechnology Engineering by Universidade Lusófona

2005-2006: Research student in Microbial and Enzyme Technology Lab at ITQB under the supervision of Prof. Lígia Oliveira Martins

Since 2006: PhD fellowship by Fundação para a Ciência e Tecnologia, in Microbial and Enzyme Technology Lab at ITQB under the supervision of Prof. Lígia Oliveira Martins