title: The ferredoxin:NAD+ oxidoreductase (Rnf) from the acetogen Acetobacterium woodii requires Na+ and is reversibly coupled to the membrane potential.
abstract: The anaerobic acetogenic bacterium Acetobacterium woodii has a novel Na(+)-translocating electron transport chain that couples electron transfer from reduced ferredoxin to NAD(+) with the generation of a primary electrochemical Na(+) potential across its cytoplasmic membrane. In previous assays in which Ti(3+) was used to reduce ferredoxin, Na(+) transport was observed, but not a Na(+) dependence of the electron transfer reaction. Here, we describe a new biological reduction system for ferredoxin in which ferredoxin is reduced with CO, catalyzed by the purified acetyl-CoA synthase/CO dehydrogenase from A. woodii. Using CO-reduced ferredoxin, NAD(+) reduction was highly specific and strictly dependent on ferredoxin and occurred at a rate of 50 milliunits/mg of protein. Most important, this assay revealed for the first time a strict Na(+) dependence of this electron transfer reaction. The Km was 0.2 mm. Na(+) could be partly substituted by Li(+). Na(+) dependence was observed at neutral and acidic pH values, indicating the exclusive use of Na(+) as a coupling ion. Electron transport from reduced ferredoxin to NAD(+) was coupled to electrogenic Na(+) transport, indicating the generation of ΔμNa(+). Vice versa, endergonic ferredoxin reduction with NADH as reductant was possible, but only in the presence of ΔμNa(+), and was accompanied by Na(+) efflux out of the vesicles. This is consistent with the hypothesis that Rnf also catalyzes ferredoxin reduction at the expense of an electrochemical Na(+) gradient. The physiological significance of this finding is discussed.
authors.affiliation: From the Department of Molecular Microbiology and Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/Main, 60438 Frankfurt, Germany.
authors.forename: Verena
authors.initials: V
authors.lastname: Hess
authors.forename: Kai
authors.initials: K
authors.lastname: Schuchmann
authors.forename: Volker
authors.initials: V
authors.lastname: Müller
mesh_terms: D045855:Acetobacterium / Q000201:enzymology*; D001426:Bacterial Proteins / Q000378:metabolism*; D002248:Carbon Monoxide / Q000378:metabolism; D005287:Ferredoxin-NADP Reductase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008094:Lithium / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology*; D010084:Oxidation-Reduction; D012964:Sodium / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D002248:Carbon Monoxide; D008094:Lithium; D012964:Sodium; D005287:Ferredoxin-NADP Reductase
keywords: Bioenergetics/Electron Transfer Complex; Electron Transfer; Energy Metabolism; Enzyme Kinetics; Membrane Energetics; Membrane Transport; Oxidation-Reduction; Physiology; Rnf; Sodium Transport
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