title: Identification of two [4Fe-4S]-cluster-containing hydro-lyases from Pyrococcus furiosus.
abstract: The hyperthermophilic archaeon Pyrococcus furiosus is a strict anaerobe. It is therefore not expected to use the oxidative tricarboxylic acid (TCA) cycle for energy transduction. Nonetheless, its genome encodes more putative TCA cycle enzymes than the closely related Pyrococcus horikoshii and Pyrococcus abyssi, including an aconitase (PF0201). Furthermore, a two-subunit fumarase (PF1755 and PF1754) is encoded on the Pyr. furiosus genome. In the present study, these three genes were heterologously overexpressed in Escherichia coli to enable characterization of the enzymes. PF1755 and PF1754 were shown to form a [4Fe-4S]-cluster-containing heterodimeric enzyme, able to catalyse the reversible hydratation of fumarate. The aconitase PF0201 also contained an Fe-S cluster, and catalysed the conversion from citrate to isocitrate. The fumarase belongs to the class of two-subunit, [4Fe-4S]-cluster-containing fumarate hydratases exemplified by MmcBC from Pelotomaculum thermopropionicum; the aconitase belongs to the aconitase A family. Aconitase probably plays a role in amino acid synthesis when the organism grows on carbohydrates. However, the function of the seemingly metabolically isolated fumarase in Pyr. furiosus has yet to be established.
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Barbara M A
authors.initials: BMA
authors.lastname: van Vugt-Lussenburg
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Laura
authors.initials: L
authors.lastname: van der Weel
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Wilfred R
authors.initials: WR
authors.lastname: Hagen
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Peter-Leon
authors.initials: PL
authors.lastname: Hagedoorn
mesh_terms: D000154:Aconitate Hydratase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D002952:Citric Acid Cycle; D004926:Escherichia coli / Q000378:metabolism; D005649:Fumarate Hydratase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D020027:Pyrococcus furiosus / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D013057:Spectrum Analysis
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D005649:Fumarate Hydratase; D000154:Aconitate Hydratase