title: The membrane-bound [NiFe]-hydrogenase (Ech) from Methanosarcina barkeri: unusual properties of the iron-sulphur clusters.
abstract: The purified membrane-bound [NiFe]-hydrogenase from Methanosarcina barkeri was studied with electron paramagnetic resonance (EPR) focusing on the properties of the iron-sulphur clusters. The EPR spectra showed signals from three different [4Fe-4S] clusters. Two of the clusters could be reduced under 101 kPa of H2, whereas the third cluster was only partially reduced. Magnetic interaction of one of the clusters with an unpaired electron localized on the Ni-Fe site indicated that this was the proximal cluster as found in all [NiFe]-hydrogenases. Hence, this cluster was assigned to be located in the EchC subunit. The other two clusters could therefore be assigned to be bound to the EchF subunit, which has two conserved four-Cys motifs for the binding of a [4Fe-4S] cluster. Redox titrations at different pH values demonstrated that the proximal cluster and one of the clusters in the EchF subunit had a pH-dependent midpoint potential. The possible relevance of these properties for the function of this proton-pumping [NiFe]-hydrogenase is discussed.
authors.affiliation: Swammerdam Institute for Life Sciences, Biochemistry, University of Amsterdam, the Netherlands; Max-Planck-Institut für Terrestrische Mikrobiologie, Marburg, Germany.
authors.forename: Sergei
authors.initials: S
authors.lastname: Kurkin
authors.forename: Jörn
authors.initials: J
authors.lastname: Meuer
authors.forename: Jürgen
authors.initials: J
authors.lastname: Koch
authors.forename: Reiner
authors.initials: R
authors.lastname: Hedderich
authors.forename: Simon P J
authors.initials: SP
authors.lastname: Albracht
mesh_terms: D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000378:metabolism; D017021:Methanosarcina barkeri / Q000201:enzymology*; D008954:Models, Biological; D008956:Models, Chemical; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry*; D011485:Protein Binding; D011522:Protons; D013696:Temperature
chemical_list: D007506:Iron-Sulfur Proteins; D011522:Protons; D010088:Oxidoreductases; C400203:Ech hydrogenase