Flavodoxin from Wolinella succinogenes
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© Springer-Verlag 1996
O R I G I N A L PA P E R
Simone Biel · Oliver Klimmek · Roland Groß · Achim Kröger
Flavodoxin from Wolinella succinogenes
Received: 15 May 1996 / Accepted: 21 June 1996
Abstract A monomeric flavoprotein (18.8 kDa) was isolated from the soluble cell fraction of Wolinella succinogenes and was identified as a flavodoxin based on its Nterminal sequence, FMN content, and redox properties. The midpoint potentials of the flavodoxin (Fld) at pH 7.5 were measured as –95 mV (Fldox/Flds) and –450 mV (Flds/Fldred) relative to the standard hydrogen electrode. The cellular flavodoxin content [0.3 µmol (g protein)–1] was the same in bacteria grown with fumarate or with polysulfide as the terminal acceptor of electron transport. The flavodoxin did not accept electrons from hydrogenase or formate dehydrogenase, the donor enzymes of electron transport to fumarate or polysulfide. Pyruvate:flavodoxin oxidoreductase activity [180 U (g cellular protein)–1] was detected in the soluble cell fraction of W. succinogenes grown with fumarate or polysulfide. The enzyme was equally active with Fldox or Flds at high concentrations. The Km for Flds (80 µM) was larger than that for Fldox and for the ferredoxin isolated from W. succinogenes (15 µM). We conclude that flavodoxin serves anabolic rather than catabolic functions in W. succinogenes. Key words Flavodoxin · Wolinella succinogenes · Pyruvate synthase Abbreviations Fldox Oxidized flavodoxin · Flds Flavodoxin radical (semiquinone) · Fldred Reduced flavodoxin · DMN 2,3-Dimethyl-1,4-naphthoquinone
Introduction Flavodoxins are a group of small, monomeric flavoproteins that contain a single molecule of FMN and function as electron carriers in low-potential redox reactions (Mayhew and Tollin 1992). Flavodoxins have been isolated from strictly anaerobic, from facultative, from obligately aerobic, and from phototrophic bacteria as well as from eucaryotic algae. Oxidized flavodoxins can accept two electrons. Depending on the species, the first electron is accepted at standard potentials ranging between +50 and –280 mV, and the second between –320 and –520 mV. Flavodoxins may function as a substrate of hydrogenase, nitrogenase, and pyruvate synthase (EC 1.2.7.1) (Mayhew and Ludwig 1975) and as a component of the electron transport to sulfite (Irie et al. 1973). Furthermore, certain enzymes (e.g., pyruvate:formate lyase; Osborne et al. 1991) are activated by electron transfer from flavodoxin. This paper describes the isolation and characterization of the flavodoxin from Wolinella succinogenes and its role in the metabolism of this anaerobic bacterium. Recent observations have suggested that a cytoplasmic redox protein such as flavodoxin might function as a mediator in the electron transport from hydrogenase or formate dehydrogenase to polysulfide reductase (Jankielewicz et al. 1995; Kotzian et al. 1996). Therefore, we tested whether flavodoxin serves as electron acceptor of hydrogenase or formate dehydrogenase.
Materials and methods Strain and culture conditions Dedicated to
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