Unique Carbon Source Research Articles

Degradation of sodium anthraquinone sulphonate by free and immobilized bacterial cultures

Aerobic biodegradation of a xenobiotic recalcitrant compound sodium anthraquinone-2-sulphonate (SAS), was investigated using as an inoculum a mixed microbial culture, which was activated sludge from industrial and domestic waste-water treatment plants. The difference in SAS degradation was examined using two main systems: (1) suspended cells and (2) immobilized cells, both in batch and in continuous culture. In the suspended cell system, under continuous culture conditions using SAS as a unique source of carbon and energy, it was possible to degrade about 95% of this substrate after 6 days. Maximal SAS removal rates in the suspended-cell system were 593 mg SAS l−1 h−1 and 88.7 mg SAS l−1 h−1 for dilution rates (D) of 0.05 h−1 and 0.075 h−1, respectively. In the immobilized-cell system, almost all SAS was degraded in 6 days and the maximal removal rate reached 88.7 mg SAS l−1 h−1 at D=0.05 h−1. Application of a continuous-flow enrichment procedure resulted in selection of several kinds of micro-organisms and led to a progressive elimination of some species of Aeromonas. A stable microbial community of 11 strains has been established and characterized at D=0.075 h−1. Most of them were Gram-negative and belonged to the genus Pseudomonas.

Non-cooperative effects of glucose and 2-deoxyglucose on their metabolism in Saccharomyces cerevisiae studied by 1H-NMR and 13C-NMR spectroscopy.

The reciprocal effects of 2-deoxy-D-glucose (dGlc) and glucose (Glc) on the aerobic metabolism of Glc and dGlc in Glc-grown repressed Saccharomyces cerevisiae were studied at 30 degrees C in a standard pyrophosphate medium containing about 4.5 x 10(7) cells/ml. 1H-, 13C-NMR and biochemical techniques were used for quantitative evaluation of Glc and dGlc metabolites. The detection of intracellular dGlc and the determination of the intracellular dGlc6P/dGlc ratio were realised using [1-13C]dGlc and 13C-NMR spectroscopy. The rates of Glc consumption in the absence and in the presence of 5 mM dGlc and 20 mM dGlc were 29 +/- 0.03 mM/min (n = 3), 16 +/- 0.02 mM/min (n = 3), and 0.08 +/- 0.01 mM/min (n = 3), respectively. This means that the Glc consumption is reduced about 41% and 70% in the presence of 5 mM and 20 mM dGlc, respectively. When dGlc is the unique carbon source, only alpha and beta anomers of dGlc6P were formed. Their quantities are equivalent and reach the maximum values within 1 h of incubation and then decrease gradually. By contrast, Glc favours the consumption of dGlc and the synthesis of dGlc6P, dideoxy-trehalose (dGlc-dGlc), deoxy-trehalose (dGlc-Glc). In the presence of Glc, dGlc6P reaches a plateau after 1 h or 2 h of incubation while the quantities of trehalose (Glc-Glc), dGlc-dGlc, dGlc-Glc, which are small at 1 h, rapidly increase with time. The reasons why dGlc and Glc exert opposite effects on their metabolism are discussed. The production of Glc-Glc decreases with increasing the external dGlc concentration or the dGlc/Glc ratio. The effect of dGlc on the biosynthesis of Glc-Glc can be explained by the competition of dGlc and Glc with respect to hexokinase. Although Glc favours the synthesis of dGlc6P, the maximum concentration of dGlc6P shows little dependence on the external dGlc concentration as long as glucose is not exhausted. The internal dGlc6P/dGlc ratio at equilibrium, about 4.7 +/- 0.7, is also found to be independent of the dGlc concentration in the suspension. Only a small fraction of dGlc6P disappears to give rise to the formation of dGlc-dGlc and dGlc-Glc. At equilibrium the inverse reaction from dGlc6P to dGlc may be important to compensate for the fast reaction of dGlc phosphorylation by hexokinase. At least nine series of experiments were conducted and showed that, in pyrophosphate media and for incubation times less than 4 h, dGlc-dGlc was not observed in the absence of Glc.(ABSTRACT TRUNCATED AT 400 WORDS)

Microbial decomposition of some cyclodextrin derivatives by bacteria associated with plants

Bacteria associated with plants were examined for their ability to use cyclodextrins (CDs) as a unique carbon source. Among 24 strains from eight genera, Xanthomonas were the most active. The cyclodextrin decomposing ability of strains were not related to their taxonomic position. Chemical structure significantly influenced the degradability of cyclodextrins by bacteria. The less rigid ring structure of gamma-CD favoured utilization, whereas methylation led to reduced utilization.

Selection of microorganisms which produce raw-starch degrading enzymes

Microorganisms which produce strong raw-starch degrading enzymes were isolated from soil using a medium containing a unique carbon source, “α-amylase resistant starch (α-RS)”, which is insoluble in water and hardly digested with Bacillus amyloliquefaciens α-amylase. Among the isolates, three strains showing high activities were characterized. Two of them, K-27 (fungus) and K-28 (yeast), produced α-amylase and glucoamylase, and the final product from starch was only glucose. The third strain, K-2, was a bacterium and produced α-amylase, which produced glucose and malto-oligosaccharides from starch. The enzyme preparation of these strains degraded raw corn starch rapidly.

Study of the biosynthesis of 3-isopropyl-2-methoxypyrazine produced by Pseudomonas taetrolens

The biosynthesis pathway of 3-isopropyl-2-methoxypyrazine (1) by Pseudomonas taetrolens was studied. Cultivation of the bacterium in a broth containing l-leucine as a unique source of carbon and nitrogen allowed an improvement of the yield of 1 which was up to 10 mg/litre. 13C- l-valine was introduced into the medium. Involvement of the amino-acid in the 1 biosynthesis could be proved by following the incorporation of the labelling by both MS and 1H and 13C-NMR. Attempts to introduce 13C-glycine (which was supposed to be the second precursor) into the 1 molecule were unsuccessful.

Preparation Microbiologique Du 2‐CETO‐3‐Desoxy‐D‐Gluconate 1‐14C ou U‐14C

AbstractA E. Coli K 12 mutant having lost 2‐keto‐3‐desoxy‐gluconate kinase activity is unable to utilize glucuronate as a unique carbon source. However, this strain is able to partially convert glucuronate into KDG in resting cells conditions reported here.A method is described making it possible to produce 1‐ 14C‐KDG or U‐14C‐KDG when the kinase cells metabolise 6‐14C or U‐14C‐D‐glucuronate. The radioactive KDG accumulated in the incubation medium has been purified thanks to a biological specific method followed by chromatography on anionic resin. The yield of the purified KDG from the hexuronate is about 20%. According to chromatographic criteria and specific conversion by KDG‐kinase, the product obtained has been authenticated as true KDG.

A common pathway for hexouronate degradation in Escherichia coli K 12. Induction mechanism of 2-keto-3-deoxy-gluconate metabolizing enzymes

After the recent investigation on the biochemical properties of the 2‐keto‐3‐deoxy‐glucono‐kinase and 2‐keto‐3‐deoxy‐6‐phospho‐gluconate aldolase in Escherichia coli K12, we report in this paper the induction mechanism of these two enzymes and the genetic control of a new enzymatic activity: the 2‐keto‐3‐deoxy‐gluconate transport system.The d‐glucuronate and d‐galacturonate raise the high basal levels of the base and aldolase on glycerol by a factor of 5 and 3–4, respectively, the kinase and aldolase being enzymes of the common degradative pathway of d‐glucuronate and d‐galacturonate. The differential rates of synthesis of these two enzymes are constant from the addition of the galacturonate. Under a variety of conditions (different inducers and substrates of growth) kinase and aldolase have been found to be synthesized non coordinately. The strongest decoordination is carried out by gluconate: this compound which is a good inducer of the aldolase, not only is unable to induce the kinase but also represses it. Moreover, the aldolase seems to be less sensitive to the catabolic repression than the kinase.By means of appropriate negative mutants we have established that the two hexuronates and two keto‐uronates do not induce directly kinase and aldolase, but by sequential conversion into 2‐keto‐3‐deoxy‐gluconate. Furthermore, the fact that the hexuronates induce the kinase in aldolase‐negative mutants (kdg A) and besides, “over” induce the aldolase in kinase negative mutants (kdg K) strongly suggests that 2‐keto‐3‐deoxy‐gluconate is a true inducer for both enzymes. In addition, the gluconate which still induces the aldolase in an edd strain suggests that, this compound and/or the 6‐phospho‐gluconate are/is also inducer(s) of this enzyme.Moreover, we have isolated spontaneous mutants able to utilize the 2‐keto‐3‐deoxy‐gluconate as a unique carbon source. Two classes have been identified. The mutants of these classes which arise at high frequency (10−5 to 10−6) are either simultaneously constitutive for a specific 2‐keto‐3‐deoxy‐gluconate transport system, kinase and aldolase (class kdg Rc)or only constitutive for the transport system (class kdg Pc). The 2‐keto‐3‐deoxy‐gluconate as exogenous substrate, which cannot induce its transport system in the wild type, behaves like a non‐inducer substrate. So the strains kdg Rc and kdg Pc are respectively i− and Oclac‐like mutants. These findings and the fact that we have found thermosensitive mutants mapping in the kdg R locus (34.5 min) and which are simultaneously derepressed for the permease, kinase and aldolase at 42°C but not at 28°C, strongly suggest that the synthesis of the three sequential enzymes degrading the 2‐keto‐3‐deoxy‐gluconate is negatively controlled by a common regulator gene product. The decoordinated syntheses of these three enzymes are in agreement with the scattering of the corresponding operons on the chromosome: transport system operon (76.5min), kinase operon (69 min), aldolase operon (34 min).