title: The iron-sulfur cluster-free hydrogenase (Hmd) is a metalloenzyme with a novel iron binding motif.
abstract: The iron-sulfur cluster-free hydrogenase (Hmd) from methanogenic archaea harbors an iron-containing cofactor of yet unknown structure. X-ray absorption spectroscopy of the active, as isolated enzyme from Methanothermobacter marburgensis (mHmd) and of the active, reconstituted enzyme from Methanocaldococcus jannaschii (jHmd) revealed the presence of mononuclear iron with two CO, one sulfur and one or two N/O in coordination distance. In jHmd, the single sulfur ligand is most probably provided by Cys176, as deduced from a comparison of the activity and of the x-ray absorption and Mössbauer spectra of the enzyme mutated in any of the three conserved cysteines. In the isolated Hmd cofactor, two CO, one sulfur, and two nitrogen/oxygen atoms coordinate the iron, the sulfur ligand being most probably provided by mercaptoethanol, which is absolutely required for the extraction of the iron-containing cofactor from the holoenzyme and for the stabilization of the extracted cofactor. In active mHmd holoenzyme, the number of iron ligands increased by one when one of the Hmd inhibitors (CO or KCN) were present, indicating that in active Hmd, the iron contains an open coordination site, which is proposed to be the site of H2 interaction.
authors.affiliation: European Molecular Biology Laboratory (EMBL), Outstation Hamburg at Deutsches Electronen Synchroton (DESY), Notkestrasse 85, D-22603 Hamburg, Germany.
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Korbas
authors.forename: Sonja
authors.initials: S
authors.lastname: Vogt
authors.forename: Wolfram
authors.initials: W
authors.lastname: Meyer-Klaucke
authors.forename: Eckhard
authors.initials: E
authors.lastname: Bill
authors.forename: Erica J
authors.initials: EJ
authors.lastname: Lyon
authors.forename: Rudolf K
authors.initials: RK
authors.lastname: Thauer
authors.forename: Seigo
authors.initials: S
authors.lastname: Shima
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry*; D001665:Binding Sites; D002244:Carbon / Q000737:chemistry; D002248:Carbon Monoxide / Q000737:chemistry; D004789:Enzyme Activation; D006864:Hydrogenase / Q000737:chemistry; D007501:Iron / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D008024:Ligands; D017013:Methanobacteriaceae / Q000201:enzymology; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D013052:Spectrometry, X-Ray Emission; D013816:Thermodynamics
chemical_list: D019843:Archaeal Proteins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D002244:Carbon; D002248:Carbon Monoxide; D007501:Iron; D006864:Hydrogenase