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V Müller – One of the best experts on this subject based on the ideXlab platform.

  • Exploring Bacterial Microcompartments in the Acetogenic Bacterium Acetobacterium woodii.
    Frontiers in microbiology, 2020
    Co-Authors: Nilanjan Pal Chowdhury, Lydia Alberti, Mark Linder, V Müller
    Abstract:

    The strictly anaerobic acetogenic bacterium Acetobacterium woodii is metabolically diverse and grows on variety of substrates which includes H2 + CO2, sugars, alcohols and diols. It is unique in producing bacterial microcompartments (BMC) during growth on different substrates such as 1,2-propanediol, 2,3-butanediol, ethanol or fructose. In this study, we analyzed the genetic organization and expression of the BMC genes within the A. woodii genome, the previously described 18 gene pdu cluster as well as four other cluster potentially encoding one or two shell proteins. Expression analysis of respective gene clusters revealed that the pdu gene cluster is highly expressed during growth on 1,2-PD, 2,3-BD, ethanol and ethylene glycol. The promoter region upstream of the pduA gene was identified and used to establish a reporter gene assay based on chloramphenicol acetyl transferase as a reporter protein. The reporter gene assay confirmed the qPCR data and demonstrated that 1,2-PD is superior over ethanol and ethylene glycol as inducer. BMCs were enriched from cells grown on 2,3- BD and 1,2-PD and shown to have typical structure in electron micrographs. Biochemical analyses revealed several of the protein encoded by the pdu cluster to be part of the isolated BMCs. These data demonstrate a very unique situation in A. woodii in which apparently one BMC gene cluster in expressed during growth on different substrates.

  • Alanine, a Novel Growth Substrate for the Acetogenic Bacterium Acetobacterium woodii
    Applied and environmental microbiology, 2018
    Co-Authors: Judith Dönig, V Müller
    Abstract:

    ABSTRACT Acetogenic bacteria are an ecophysiologically important group of strictly anaerobic bacteria that grow lithotrophically on H2 plus CO2 or on CO or heterotrophically on different substrates such as sugars, alcohols, aldehydes, or acids. Amino acids are rarely used. Here, we describe that the model acetogen Acetobacterium woodii can use alanine as the sole carbon and energy source, which is in contrast to the description of the type strain. The alanine degradation genes have been identified and characterized. A key to alanine degradation is an alanine dehydrogenase which has been characterized biochemically. The resulting pyruvate is further degraded to acetate by the known pathways involving the Wood-Ljungdahl pathway. Our studies culminate in a metabolic and bioenergetic scheme for alanine-dependent acetogenesis in A. woodii. IMPORTANCE Peptides and amino acids are widespread in nature, but there are only a few reports that demonstrated use of amino acids as carbon and energy sources by acetogenic bacteria, a central and important group in the anaerobic food web. Our finding that A. woodii can perform alanine oxidation coupled to reduction of carbon dioxide not only increases the number of substrates that can be used by this model acetogen but also raises the possibility that other acetogens may also be able to use alanine. Indeed, the alanine genes are also present in at least two more acetogens, for which growth on alanine has not been reported so far. Alanine may be a promising substrate for industrial fermentations, since acid formation goes along with the production of a base (NH3) and pH regulation is a minor issue.

  • Regulation of lactate metabolism in the acetogenic bacterium Acetobacterium woodii.
    Environmental microbiology, 2018
    Co-Authors: Marie Charlotte Schoelmerich, Alexander Katsyv, Woung Sung, Vanessa Mijic, Anja Wiechmann, Patrick Kottenhahn, Jonathan Baker, Nigel P. Minton, V Müller
    Abstract:

    Acetogenic bacteria compete in an energy-limited environment by coupling different metabolic routes to their central metabolism of CO2 fixation. The underlying regulatory mechanisms are often still not understood. In this work, we analysed how lactate metabolism is regulated in the model acetogen Acetobacterium woodii. Construction of a ΔlctCDEF mutant and growth analyses demonstrated that the genes are essential for growth on lactate. Subsequent bridging PCR and quantitative PCR analyses revealed that the lctBCDEF genes form an operon that was expressed only during lactate metabolism. The lctA gene was cloned, expressed in Escherichia coli and purified. LctA bound to the intergenic DNA region between lctA and the lct operon in electromobility shift assays, and binding was revoked in the presence of lactate. Further restriction site protection analyses consolidated the lactate-dependent binding of LctA and identified the binding site within the DNA. Cells grew mixotrophically on lactate and another energy source and showed no diauxic growth. From these data, we conclude that the catabolic lactate metabolism is encoded by the lct operon and its expression is negatively regulated by the DNA-binding repressor LctA.

Jutta Reidlinger – One of the best experts on this subject based on the ideXlab platform.

  • The molecular structure of the Na+‐translocating F1F0‐ATPase of Acetobacterium woodii, as revealed by electron microscopy, resembles that of H+‐translocating ATPases
    FEBS letters, 1994
    Co-Authors: Jutta Reidlinger, Frank Mayer, V Müller
    Abstract:

    The Na+-translocating F1F0-ATPase of Acetobacterium woodii was examined by electron microscopy. After reconstitution into proteoliposomes, knobs typical for the F1 domain were visible on the outside of the membrane. The F1-part of the isolated enzyme showed a hexagonal symmetry suggesting an α3β3 structure, and the F1F0 complex had molecular dimensions very similar to those of H+-translocating ATPases of E. coli, chloroplasts, and mitochondria.

  • the molecular structure of the na translocating f1f0 atpase of Acetobacterium woodii as revealed by electron microscopy resembles that of h translocating atpases
    FEBS Letters, 1994
    Co-Authors: Jutta Reidlinger, Frank Mayer, V Müller
    Abstract:

    The Na+-translocating F1F0-ATPase of Acetobacterium woodii was examined by electron microscopy. After reconstitution into proteoliposomes, knobs typical for the F1 domain were visible on the outside of the membrane. The F1-part of the isolated enzyme showed a hexagonal symmetry suggesting an α3β3 structure, and the F1F0 complex had molecular dimensions very similar to those of H+-translocating ATPases of E. coli, chloroplasts, and mitochondria.

  • Purification of ATP synthase from Acetobacterium woodii and identification as a Na+‐translocating F1FO‐type enzyme
    European journal of biochemistry, 1994
    Co-Authors: Jutta Reidlinger, V Müller
    Abstract:

    The ATPase of Acetobacterium woodii was purified after solubilization of membranes with Triton X-100 by poly(ethylene glycol) precipitation and gel filtfiltration. The enzyme consists of at least six subunits of apparent molecular masses of 57, 52, 35, 19, 15 and 4.8 kDa, as determined by SDS/PAGE. The 52-kDa band is immunologically related to the F1F0-ATPase beta subunit of Escherichia coli. The enzyme is not inhibited by vanadate but is inhibited by nitrate, azide and N,N’-dicyclohexylcarbodiimide; the 4.8-kDa subunit specifically reacts with N,N’-dicyclohexyl[14C]carbodiimide, indicating that the enzyme is of the F1F0 type. The enzyme activity is dependent on MgATP (Km = 0.4), has a pH optimum of pH 7-9 and is stimulated by sulfite. ATP hydrolysis is strictly dependent on sodium ions with a Km for Na+ of 0.4 mM. The purified enzyme was reconstituted into liposomes. Upon addition of ATP, primary and electrogenic 22Na+ transport into the lumen of the proteoliposomes was determined. These experiments demonstrate that the ATPase of Acetobacterium woodii is a Na(+)-translocating F1F0-type ATPase.

S Rahlfs – One of the best experts on this subject based on the ideXlab platform.

  • sequence of subunit a of the na translocating f1f0 atpase of Acetobacterium woodii proposal for residues involved in na binding
    FEBS Letters, 1999
    Co-Authors: S Rahlfs, V Müller
    Abstract:

    Na+ transport through the F0 domain of Na+-F1F0-ATPases involves the combined action of subunits c and a but the residues involved in Na+ liganding in subunit a are unknown. As a first step towards the identification of these residues, we have cloned and sequenced the gene encoding subunit a of the Na+-F1F0-ATPase of Acetobacterium woodii. This is the second sequence available now for this subunit from Na+-F1F0-ATPases. A comparison of subunit a from Na+-F1F0-ATPases with those from H+-translocating enzymes unraveled structural similarity in a C-terminal segment including the ultimate and penultimate transmembrane helix. Seven residues are conserved in this region and, therefore, likely to be involved in Na+ liganding.

  • Sequence of subunit a of the Na(+)-translocating F1F0-ATPase of Acetobacterium woodii: proposal for residues involved in Na+ binding.
    FEBS Letters, 1999
    Co-Authors: S Rahlfs, V Müller
    Abstract:

    Na+ transport through the F0 domain of Na+-F1F0-ATPases involves the combined action of subunits c and a but the residues involved in Na+ liganding in subunit a are unknown. As a first step towards the identification of these residues, we have cloned and sequenced the gene encoding subunit a of the Na+-F1F0-ATPase of Acetobacterium woodii. This is the second sequence available now for this subunit from Na+-F1F0-ATPases. A comparison of subunit a from Na+-F1F0-ATPases with those from H+-translocating enzymes unraveled structural similarity in a C-terminal segment including the ultimate and penultimate transmembrane helix. Seven residues are conserved in this region and, therefore, likely to be involved in Na+ liganding.

  • Sequence of subunit c of the Na(+)-translocating F1F0 ATPase of Acetobacterium woodii: proposal for determinants of Na+ specificity as revealed by sequence comparisons.
    FEBS letters, 1997
    Co-Authors: S Rahlfs, V Müller
    Abstract:

    A 3.2 kb EcoRI fragment carrying genes for Na(+)-F1F0 ATPase was cloned from chromosomal DNA of Acetobacterium woodii. DNA sequence analysis revealed the presence of an open reading frame which was identified by data base searches and comparison with the experimentally derived N-terminal amino acid sequence to code for subunit c of Na(+)-F1F0 ATPase. A comparison of the primary sequences of the two well established Na(+)-translocating F1F0 ATPases from Acetobacterium woodii and Propionigenium modestum with H(+)-translocating enzymes indicates the length of the C-terminus as well as specific residues located in the cytoplasmic membrane to be important for Na+ transport.

Eva Biegel – One of the best experts on this subject based on the ideXlab platform.

  • a na translocating pyrophosphatase in the acetogenic bacterium Acetobacterium woodii
    Journal of Biological Chemistry, 2011
    Co-Authors: Eva Biegel, V Müller
    Abstract:

    The anaerobic acetogenic bacterium Acetobacterium woodii employs a novel type of Na+-motive anaerobic respiration, caffeate respiration. However, this respiration is at the thermodynamic limit of energy conservation, and even worse, in the first step, caffeate is activated by caffeyl-CoA synthetase, which hydrolyzes ATP to AMP and pyrophosphate. Here, we have addressed whether or not the energy stored in the anhydride bond of pyrophosphate is conserved by A. woodii. Inverted membrane vesicles of A. woodii have a membrane-bound pyrophosphatase that catalyzes pyrophosphate hydrolysis at a rate of 70–120 milliunits/mg of protein. Pyrophosphatase activity was dependent on the divalent cation Mg2+. In addition, activity was strictly dependent on Na+ with a Km of 1.1 mm. Hydrolysis of pyrophosphate was accompanied by 22Na+ transport into the lumen of the inverted membrane vesicles. Inhibitor studies revealed that 22Na+ transport was primary and electrogenic. Next to the Na+-motive ferredoxin:NAD+ oxidoreductase (Fno or Rnf), the Na+-pyrophosphatase is the second primary Na+-translocating enzyme in A. woodii.

  • A Na+-translocating pyrophosphatase in the acetogenic bacterium Acetobacterium woodii.
    The Journal of biological chemistry, 2010
    Co-Authors: Eva Biegel, V Müller
    Abstract:

    The anaerobic acetogenic bacterium Acetobacterium woodii employs a novel type of Na+-motive anaerobic respiration, caffeate respiration. However, this respiration is at the thermodynamic limit of energy conservation, and even worse, in the first step, caffeate is activated by caffeyl-CoA synthetase, which hydrolyzes ATP to AMP and pyrophosphate. Here, we have addressed whether or not the energy stored in the anhydride bond of pyrophosphate is conserved by A. woodii. Inverted membrane vesicles of A. woodii have a membrane-bound pyrophosphatase that catalyzes pyrophosphate hydrolysis at a rate of 70–120 milliunits/mg of protein. Pyrophosphatase activity was dependent on the divalent cation Mg2+. In addition, activity was strictly dependent on Na+ with a Km of 1.1 mm. Hydrolysis of pyrophosphate was accompanied by 22Na+ transport into the lumen of the inverted membrane vesicles. Inhibitor studies revealed that 22Na+ transport was primary and electrogenic. Next to the Na+-motive ferredoxin:NAD+ oxidoreductase (Fno or Rnf), the Na+-pyrophosphatase is the second primary Na+-translocating enzyme in A. woodii.

  • the ins and outs of na bioenergetics in Acetobacterium woodii
    Biochimica et Biophysica Acta, 2009
    Co-Authors: Silke Schmidt, Eva Biegel, V Müller
    Abstract:

    Abstract The acetogenic bacterium Acetobacterium woodii uses a transmembrane electrochemical sodium ion potential for bioenergetic reactions. A primary sodium ion potential is established during carbonate (acetogenesis) as well as caffeate respiration. The electrogenic Na+ pump connected to the Wood–Ljungdahl pathway (acetogenesis) still remains to be identified. The pathway of caffeate reduction with hydrogen as electron donor was investigated and the only membrane-bound activity was found to be a ferredoxin-dependent NAD+ reduction. This exergonic electron transfer reaction may be catalyzed by the membrane-bound Rnf complex that was discovered recently and is suggested to couple exergonic electron transfer from ferredoxin to NAD+ to the vectorial transport of Na+ across the cytoplasmic membrane. Rnf may also be involved in acetogenesis. The electrochemical sodium ion potential thus generated is used to drive endergonic reactions such as flagellar rotation and ATP synthesis. The ATP synthase is a member of the F1FO class of enzymes but has an unusual and exceptional feature. Its membrane-embedded rotor is a hybrid made of FO and VO-like subunits in a stoichiometry of 9:1. This stoichiometry is apparently not variable with the growth conditions. The structure and function of the Rnf complex and the Na+ F1FO ATP synthase as key elements of the Na+ cycle in A. woodii are discussed.

Gerhard Gottschalk – One of the best experts on this subject based on the ideXlab platform.

  • Acetogenesis and ATP synthesis in Acetobacterium itwoodii are coupled via a transmembrane primary sodium ion gradient
    FEMS Microbiology Letters, 1993
    Co-Authors: Reno Heise, V Müller, Gerhard Gottschalk
    Abstract:

    In cell suspensions of Acetobacterium woodii the acetyl-CoA pathway is coupled to net ATP formation. Acetate formation as well as ATP synthesis and the generation of a transmembrane sodium ion gradient are not inhibited by protonophores but by sodium ionophores. Acetogenesis from CO or formaldehyde + CO as catalyzed by inverted vesicles is coupled to sodium ion uptake. Both processes are not inhibited by protonophores but by sodium ionophores. These experiments are in accordance with the presence of a primary sodium ion<ion pump connected to the acetyl-CoA pathway which enables the cells to synthesize net ATP by means of a ΔμNa+ in concert with a Na+-translocating ATPase.

  • Presence of a sodium-translocating ATPase in membrane vesicles of the homoacetogenic bacterium Acetobacterium woodii
    European journal of biochemistry, 1992
    Co-Authors: Reno Heise, V Müller, Gerhard Gottschalk
    Abstract:

    Inverted membrane vesicles of the homoacetogenic bacterium Acetobacterium woodii catalyzed the hydrolysis of ATP with a rate of 100–150 nmol · min−1· mg protein−1. The ATPase was stimulated 1.4–1.6-fold by NaCl and inhibited by N,N′-dicyclohexylcarbodiimide, tributyltin or azide. The degree of inhibition caused by F0-directed but not F1-directed inhibitors was affected by the Na+ concentration in the medium. These experiments indicated the presence of a sodium-translocating ATPase. This was verified by transport studies. Upon addition of ATP to inverted vesicles, 22Na+ was actively transported into the intravesicular space up to a 24-fold accumulation. Na+ transport was inhibited by the sodium ionophore N,N,N′,N′,-tetracyclohexyl-1,2-phenyl-enedioxydiacetamide but stimulated by valinomycin with potassium whereas the protonophore 3,5,-di-tert-butyl-4-hydroxybenzylidenemalonitrile was without effect. N,N′-dicyclohexylcarbodiimide and tributyltin inhibited 22Na+ transport. These experiments are in accordance with a primary electrogenic Na+ transport as catalyzed by a F1F0-ATPase.

  • A sodium-stimulated ATP synthase in the acetogenic bacterium Acetobacterium woodii
    FEBS letters, 1991
    Co-Authors: Reno Heise, V Müller, Jutta Reidlinger, Gerhard Gottschalk
    Abstract:

    Experiments with resting cells of Acetobacterium woodii were performed to elucidate the coupling ion used by the ATP synthase. A. woodii synthesized ATP in response to an artificial ΔpH, indicating the presence of a proton-translocating ATPase. On the other hand, a ΔpNa, as well as a proton diffusion potential, could serve as a driving force for ATP synthesis with the latter strictly dependent on Na+. These results are indicative for the presence of a Na+-translocating ATP synthase in A. woodii.