title: NADP-specific electron-bifurcating [FeFe]-hydrogenase in a functional complex with formate dehydrogenase in Clostridium autoethanogenum grown on CO.
abstract: Flavin-based electron bifurcation is a recently discovered mechanism of coupling endergonic to exergonic redox reactions in the cytoplasm of anaerobic bacteria and archaea. Among the five electron-bifurcating enzyme complexes characterized to date, one is a heteromeric ferredoxin- and NAD-dependent [FeFe]-hydrogenase. We report here a novel electron-bifurcating [FeFe]-hydrogenase that is NADP rather than NAD specific and forms a complex with a formate dehydrogenase. The complex was found in high concentrations (6% of the cytoplasmic proteins) in the acetogenic Clostridium autoethanogenum autotrophically grown on CO, which was fermented to acetate, ethanol, and 2,3-butanediol. The purified complex was composed of seven different subunits. As predicted from the sequence of the encoding clustered genes (fdhA/hytA-E) and from chemical analyses, the 78.8-kDa subunit (FdhA) is a selenocysteine- and tungsten-containing formate dehydrogenase, the 65.5-kDa subunit (HytB) is an iron-sulfur flavin mononucleotide protein harboring the NADP binding site, the 51.4-kDa subunit (HytA) is the [FeFe]-hydrogenase proper, and the 18.1-kDa (HytC), 28.6-kDa (HytD), 19.9-kDa (HytE1), and 20.1-kDa (HytE2) subunits are iron-sulfur proteins. The complex catalyzed both the reversible coupled reduction of ferredoxin and NADP(+) with H2 or formate and the reversible formation of H2 and CO2 from formate. We propose the complex to have two functions in vivo, namely, to normally catalyze CO2 reduction to formate with NADPH and reduced ferredoxin in the Wood-Ljungdahl pathway and to catalyze H2 formation from NADPH and reduced ferredoxin when these redox mediators get too reduced during unbalanced growth of C. autoethanogenum on CO (E0' = -520 mV).
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Marburg, Germany.
authors.forename: Shuning
authors.initials: S
authors.lastname: Wang
authors.forename: Haiyan
authors.initials: H
authors.lastname: Huang
authors.forename: Jörg
authors.initials: J
authors.lastname: Kahnt
authors.forename: Alexander P
authors.initials: AP
authors.lastname: Mueller
authors.forename: Michael
authors.initials: M
authors.lastname: Köpke
authors.forename: Rudolf K
authors.initials: RK
authors.lastname: Thauer
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001431:Bacteriological Techniques; D002248:Carbon Monoxide / Q000378:metabolism*; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D004579:Electron Transport; D005560:Formate Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic; D006864:Hydrogenase / Q000235:genetics / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D009249:NADP; D002248:Carbon Monoxide; D007501:Iron; C118830:iron hydrogenase; D006864:Hydrogenase; D005560:Formate Dehydrogenases
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