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Báo cáo Y học: The membrane-bound [NiFe]-hydrogenase (Ech) from Methanosarcina barkeri : unusual properties of the iron-sulphur clusters

The purified membrane-bound [NiFe]-hydrogenase from Methanosarcina barkeriwas studied with electron para-magnetic resonance (EPR) focusing on the properties of the iron–sulphur clusters. TheEPRspectra showed signals from threedifferent [4Fe)4S] clusters.Twoof the clusters couldbe reduced under 101 kPa of H2, whereas the third cluster was only partially reduced. | Eur. J. Biochem. 269 6101-6111 2002 FEBS 2002 doi 10.1046 j.1432-1033.2002.03328.x The membrane-bound NiFe -hydrogenase Ech from Methanosarcina barkeri unusual properties of the iron-sulphur clusters Sergei Kurkin1 Jorn Meuer2 Jurgen Koch2 Reiner Hedderich2 and Simon P. J. Albracht1 1 Swammerdam Institute for Life Sciences Biochemistry University of Amsterdam the Netherlands 2Max-Planck-Institut fur Terrestrische Mikrobiologie Marburg Germany 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. Keywords Ech hydrogenase iron-sulphur pH dependence redox properties. Hydrogenases catalyse the simplest chemical reaction in nature H2 2H 2e-. They are found in wide variety of microorganisms. Hydrogenases enable some organisms to use H2 as a source of reducing equivalents under both aerobic and anaerobic conditions. In other organisms the enzyme is used to reduce protons to H2 thereby releasing the reducing equivalents obtained from the anaerobic degradation of organic substrates 1 2 . On basis of