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  • 1
    ISSN: 1432-072X
    Keywords: d-Ribulose 1,5-diphosphate carboxylase ; Oxygenase activity ; Quaternary structure ; Electron microscopy ; Alcaligenes eutrophus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract d-Ribulose 1,5-diphosphate carboxylase has been purified from autotrophically grown cells of the facultative chemolithotrophic hydrogen bacteriumAlcaligenes eutrophus. The enzyme was homogeneous by the criteria of polyacrylamide gel electrophoresis. The molecular weight of the enzyme was 505000 determined by gel filtration and sucrose density gradient centrifugation, and a sedimentation coefficient of 18.2 S was obtained. It was demonstrated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis that the enzyme consists of two types of subunits of molecular weight 52000 and 13000. Electron microscopy on the intact and the partially dissociated enzyme lead to the construction of a model for the quaternary structure of the enzyme which is composed of 8 large and 8 small subunits. The most probable symmetry of the enzyme molecule is 4:2:2. Michaelis constant (K m ) values for ribulose 1,5-diphosphate, Mg2-, and CO2 were 0.59 mM, 0.33 mM, and 0.066 mM measured under air. Oxygen was a competitive inhibitor with respect to CO2 suggesting that the enzyme also exhibits an oxygenase activity. The oxygenolytic cleavage of ribulose 1,5-diphosphate was shown and a 1:1 stoichiometry between oxygen consumption and 3-phosphoglycerate formation observed.
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 112 (1977), S. 239-246 
    ISSN: 1432-072X
    Keywords: Hydrogen bacteria ; Alcaligenes eutrophus H 16 ; Leucine biosynthesis ; α-Isopropylmalate synthase ; Temperature anomaly ; Cold lability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract α-Isopropylmalate (IPM) synthase, the first enzyme in the biosynthesis of l-leucine, was purified to a specific activity of 12 μmole/min x mg protein from the valine-isoleucine double auxotrophic mutant A-81 of the hydrogen bacterium Alcaligenes eutrophus H 16. The activity in crude extracts of derepressed cells was 0.106 μmoles of isopropylmalate formed per min and per mg protein. Gel electrophoresis and regel electrophoresis of the isolated main band resulted in several distinct bands, which were not altered by the additions of substrate α-ketoisovalerate, feedback inhibitor leucine or other effectors. The isoelectric points of the enzyme protein was between 3.9 and 4.0. The molecular weight was 114500 daltons and 100000 respectively in the absence and presence of the feedback inhibitor leucine. The enzyme activity depended strongly on the pH, the optimum is at pH 8.2. The enzyme was could labile and exhibits temperature anomalies.
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 100 (1974), S. 1-4 
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
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  • 4
    ISSN: 1432-072X
    Keywords: Coryneform Hydrogen Bacterium ; Autotrophic Growth ; Entner-Doudoroff Pathway ; Hydrogenase ; Slime Formation ; Corynebacterium autotrophicum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract 1. Corynebacterium autotrophicum strain 7 C was isolated from an enrichment culture designed for propane oxidizing bacteria. The cells are Grampositive, immotile, short, irregularly formed rods. The colonies are yellow-pigmented and slimy. The yellow pigmentation is due to carotenoids. 2. Growth occurs either autotrophically in mineral medium under an atmosphere of 70% H2+20% O2+10% CO2 or heterotrophically with fructose or many organic acids as substrates. 3. The hexoses and gluconate are degraded via the Entner-Doudoroff pathway. 6-Phosphogluconate dehydrogenase is not detectable. 4. A NAD reducing hydrogenase has not been detected; the hydrogenase is localized in the particle fraction of the crude extract and reduces methylene blue. The specific activity of hydrogenase in the crude extract of autotrophically grown cells is 2400 μl H2/mg protein · hr. During growth on fructose the enzyme is constitutively formed (1200 μl H2/mg protein · hr). 5. The utilization of fructose was suppressed by hydrogen. The inhibitory effect was significant, when either fully adapted or autotrophically grown cells were exposed to a hydrogen containing atmosphere.
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  • 5
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Zusammenfassung 1. Die Bildung der zum Fructoseabbau über den Entner-Doudoroff-Weg dienenden Enzyme wird in Hydrogenomonas H 16 durch molekularen Wasserstoff reprimiert. In volladaptierten, auf Fructose gewachsenen Zellen wird der Fructoseabbau durch H2 gehemmt. Die durch H2 bewirkte Hemmung greift wahrscheinlich an der Glucose-6-phosphat-Dehydrogenase an. 2. Die aus Hydrogenomonas H 16 100fach angereicherte Glucose-6-phosphat-Dehydrogenase unterscheidet sich vom Hefeenzym durch ihre geringe Substrataffinität, durch die Fähigkeit sowohl mit NADP als auch mit NAD als Coenzym zu reagieren und durch den sigmoiden Verlauf der Substratsättigungskurve (G-6-P). 3. Das Enzym wird durch ATP und NADH2 gehemmt. Durch ATP wird die Sigmoidität der Substratsättigungskurve verstärkt. Aus den gewonnenen kinetischen Daten ist zu schließen, a) daß die durch NADH2 bewirkte Hemmung vor allem auf einer Konkurrenzreaktion zwischen NADP und NADH2 um die Bindungsstelle des Coenzyms am Enzym beruht, b) daß G-6-P-DH ein allosterisches Enzym ist, das aus mindestens vier Untereinheiten besteht und für welches ATP ein negativer allosterischer Effector ist; c) daß die Substratbindungsstellen für G-6-P der Enzymuntereinheiten kooperative Wechselwirkungen aufeinander ausüben; d) daß G-6-P-DH ein “K-System”, ist, d. h. daß der allosterische Effector den apparenten K m -Wert,jedoch nicht die maximale Geschwindigkeit (v max) der Enzymfunktion verändert. 5. G-6-P-DH aus H 16 wird durch Magnesium-Ionen nicht aktiviert. Hohe MgSO4-Konzentrationen hemmen das Enzym sogar. Durch Zusatz von MgSO4 kann die “ATP-Hemmung” vollkommen aufgehoben werden, wenn G-6-P in hoher Konzentration im Reaktionsgemisch vorliegt. Die Hemmung wird minimal, wenn das Verhältnis Mg/ATP den Wert 1,3 übersteigt.
    Notes: Summary 1. In Hydrogenomonas H 16 the synthesis of the enzymes of the Entner-Doudoroff (ED)1 degradative pathway is repressed by molecular hydrogen. In fully induced cells grown on fructose the utilization of fructose is inhibited. This inhibition results in a decrease of the rates of growth of formation of poly-β-hydroxybutyric acid by 80%. This inhibition exerted in vivo by hydrogen is probably due to an inhibition of glucose-6-phosphate (G-6-P) dehydrogenase by metabolites. 2. G-6-P-dehydrogenase has been purified from extracts of Hydrogenomonas H 16 ca. 100-fold. It differs from the yeast enzyme by a low substrate affinity, by a specificity for both NADP and NAD and by a sigmoid substrate saturation curve (G-6-P). 3. The enzyme is inhibited by ATP and NADH2. The sigmoidity of the substrate saturation curve (G-6-P) is increased by ATP. From kinetic data the following conclusions may be drawn: a) the inhibition by NADH2 is mainly caused by competition between NADP and NADH2 for the binding site of the coenzymes at the enzyme; b) G-6-P-dehydrogenase is an allosteric enzyme which is composed of at least four subunits; ATP is a negative allosteric effector; c) the binding sites of the subunits for the substrate show cooperative interactions; d) the enzyme fits the “K-system”, i.e. the allosteric effector changes the apparent K m -value without influencing the maximal velocity (v max) of enzyme action. 4. The enzyme is more or less strongly inhibited by all nucleoside triphosphates tested as well as by ADP. The inhibitory effect of AMP and other nucleoside monophosphates is negligibly low. A positive effector has not been found. 5. The enzyme from H 16 is not activated by magnesium ions. High magnesium concentrations are even inhibitory. The inhibition caused by ATP can be completely releaved by magnesium sulfate, if G-6-P is present in quasi saturating concentration. The inhibitory effect becomes minimal when the ratio Mg/ATP exceeds 1.3.
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  • 6
    ISSN: 1432-072X
    Keywords: Key words Soluble NAD-reducing hydrogenase ; Membrane-bound hydrogenase ; Diaphorase ; Rhodococcus opacus 1b ; Alcaligenes eutrophus H16
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Six new strains of Alcaligenes enriched for and isolated as nickel-resistant bacteria resemble Alcaligenes eutrophus H16 and contain both an NAD-reducing, tetrameric soluble hydrogenase and a membrane-bound hydrogenase. None of the soluble hydrogenases share with the Rhodococcus opacus MR11 enzyme tetramer the property of being cleaved easily into two dimeric moieties [a hydrogenase (βδ) and an NADH:acceptor oxidoreductase (αγ)], in the absence of nickel or at low ionic strength. The soluble hydrogenase of the newly isolated strain MR22 of R. opacus equalled that of strain MR11. The absence of a membrane-bound hydrogenase in Alcaligenes denitrificans strain 4a-2 and in Alcaligenes ruhlandii was confirmed.
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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 38 (1960), S. 52-54 
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Zusammenfassung Zur Radioaktivitätsmessung von einzelligen Mikroorganismen wird ein Verfahren unter Verwendung von Membranfiltern beschrieben. Die Korrektionskurve für die Selbstabsorption wird angegeben.
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  • 8
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Zusammenfassung Es wird ein Verfahren zur Submerskultur von Knallgasbakterien beschrieben. Es beruht auf der kräftigen Magnetrührung der Nährlösung unter einem Gemisch von H2, O2 und CO2. Der hohen O2-Empfindlichkeit der Zellen wird durch „Gradientenbegasung” Rechnung getragen. Der fakultativ chemolithotrophe Hydrogenomonas-Stamm 20 wurde bakteriologisch charakterisiert und wachstumsphysiologisch untersucht. Die Generationszeit beträgt während der log-Phase 21/6 Std, die scheinbare Verdoppelungszeit 31/5 Std (28° C).
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 46 (1963), S. 79-95 
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Summary The following early labeled compounds resulted from 14CO2-fixation by chemolithotrophically grown washed cells of Hydrogenomonas strain H 16 under hydrogen-oxygen-atmosphere: 3-phosphoglycerate, acetate and malate as well as other phosphate-esters and organic acids. After 34 secs of 14CO2-fixation additionally the following compounds were radiochromatographically identified: pentose-, hexose- and heptulose-phosphates, phosphoglycolate, citrate, fumarate, succinate, lactate, and glutamate. Under the conditions of PHBS accumulation CO2 appears to be fixed predominantly via the reactions of the reductive pentosephosphate-cycle; malate is possibly the endproduct of a second CO2-fixation. Under air with succinate as a substrate the same suspensions fixed CO2 at a lower rate (13%), however following the same path. During fixation under heterotrophic conditions radiocarbon was found to accumulate in succinate, α-ketoglutarate and malate. Organotrophically grown cells fixed carbon dioxide by the same reactive mechanism, although at a very low rate (3%).
    Notes: Zusammenfassung Als frühzeitig markierte Verbindungen der 14CO2-Fixierung durch chemolithotroph gewachsene, gewaschene Zellen von Hydrogenomonas Stamm H 16 unter Knallgasatmosphäre wurden 3-Phosphoglycerinsäure, Essigsäure und Äpfelsäure sowie weitere Phosphatester und organische Säuren nachgewiesen. Nach 34 sec 14CO2-Einbau ließen sich radioautographisch außerdem Pentose-, Hexose- und Heptulosephosphate, Phosphoglykolsäure, Citronensäure, Fumarsäure, Bernsteinsäure, Milchsäure, Glutaminsäure und andere Verbindungen identifizieren. Unter den Bedingungen der PHBS-Speicherung scheint Kohlendioxyd vorwiegend über die Reaktionen des reduktiven Pentosephosphatcyclus fixiert zu werden; Äpfelsäure ist wahrscheinlich das Produkt einer zweiten Fixierungsreaktion. Unter Luft mit Bernsteinsäure als Substrat fixierten dieselben Suspensionen 14CO2 zwar mit geringerer Geschwindigkeit (13%), jedoch zunächst über dieselben Reaktionswege. Im Laufe der Fixierung reicherte sich 14C in Bernsteinsäure, α-Ketoglutarsäure und Äpfelsäure an. Auch organotroph gewachsene Zellen fixierten CO2 auf demselben Reaktionsweg, wenn auch mit sehr geringer Rate (3%).
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 46 (1963), S. 117-126 
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Summary Isolation and properties of a new carotenoid are described. It was found in Chromatium okenii as the only carotenoid in a remarkably high concentration (50 μg/mg protein) and is characterized by the presence of a conjugated keto-group and a tertiary methoxyl-group. We suggest to name the new carotenoid okenone. Phytofluene, ξ-carotene, neurosporene and lycopene were found only in cells grown in the presence of diphenylamine.
    Notes: Zusammenfassung Es werden Isolierung und Eigenschaften eines neuen Carotinoids beschrieben. Es liegt in Chromatium okenii als einziges Carotinoid vor, ist in relativ sehr hoher Konzentration (50 μg/mg Protein) enthalten und durch eine konjugierte Ketogruppe und eine tertiäre Methoxylgruppe charakterisiert. Es wird der Name “Okenon” vorgeschlagen. Phytofluen, ξ-Carotin, Neurosporin und Lycopin wurden als Begleitcarotinoide lediglich in Zellen nachgewiesen, die in Gegenwart von Diphenylamin gewachsen waren.
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