Interrelationships in trace-element metabolism in metal toxicities in a cobalt-resistant strain of Neurospora crassa

A strain of Neurospora crassa was isolated by training the mould to grow on media containing high concentrations of Co(2+). This strain, the Co(R) strain, exhibited approximately tenfold the resistance of the parent strain to Co(2+) and Ni(2+) but not to Zn(2+) or Cu(2+). Co(2+) toxicity in the Co(R) strain was reversed by Mg(2+) but not by Fe(3+). Also, Co(2+) did not affect iron metabolism in this strain. It is suggested that the mechanism of resistance in the Co(R) strain involves an alteration in the pattern of iron metabolism such that the latter is no longer adversely affected by toxic concentrations of Co(2+). The Co(R) strain is genetically stable and is most probably a result of a resistance mutation in N. crassa induced by Co(2+).     

Studies on polyol metabolism in Aspergillus niger

Summary Sorbitol dehydrogenase has been purified about 26 fold from a strain of Aspergillus niger, growing on sorbitol as the sole source of carbon. An absolute specificity of this enzyme for sorbitol, fructose, NAD and NADH was observed. The K_m for sorbitol and fructose were found to be 9.8x10^-5 M and 6.6x10^-4 M respectively. The enzyme was inhibited by pCMB, NaF and other metal ions studied. The enzyme was slightly activated by Fe⁺⁺⁺.Part of this work was presented at the All India Conference of Microbiologists held at Baroda, 1968/69.     

Interrelationships in trace-element metabolism in metal toxicities in nickel-resistant strains of Neurospora crassa.

Three different Ni2+-resistant strains of Neurospora crassa (NiR1, NiR2 and NiR3) have been isolated. All are stable mutants and are fourfold more resistant to Ni2+ than the parent wild-type strain. NiR1 and NiR2 are also sixfold more resistant to Co2+, whereas NiR3 is only twice as resistant to Co2+; the former two are also twofold more resistant to Zn2+, but NiR3 is not. These three strains also differ in sensitivity to Cu2+. Toxicities and concomitant accumulation patterns of Ni2+, Co2+ and Cu2+ have been examined in these strains. NiR1 and NiR2, despite quantitative individual differences, generally accumulate very high amounts of Ni2+ and Co2+, and Mg2+ reverses the toxicities of these two ions by different mechanisms; Ni2+ uptake is suppressed, but not that of Co2+. In NiR3, Mg2+ controls uptake of both Ni2+ and Co2+. Studies indicate that two kinds of Ni2+-resistant strains of N. crassa exist; one kind is resistant because it can tolerate high intracellular concentrations of heavy-metal ions, whereas the other is resistant because it can control metal-ion accumulation.     

Genetic analysis of benzoate metabolism in Aspergillus niger

Summary The benzoate metabolism of Aspergillus niger was studied as part of a design to clone the benzoate-4-hydroxylase gene of this fungus on the basis of complementation. Filtration enrichment techniques yielded mutants defective for different steps of benzoate degradation: bph (benzoate-4-hydroxylase), phh (4-hydroxybenzoate-3-hydroxylase) and prc (protocatechuate ring cleavage) mutants. In this way the degradation pathway for benzoate, involving the formation of 4-hydroxybenzoate and 3,4-dihydroxybenzoate has been confirmed. In addition a mutant sensitive to benzoate has been found. Complementation tests in somatic diploids showed that the bph mutants belonged to two complementation groups. The major group is probably defective in the structural gene (bphA). All phh mutants tested belonged to one complementation group. The prc mutants could be divided into several groups on the basis of their growth on different aromatic substrates and on the basis of the complementation test. The phh and both bph mutations are shown to be located on different chromosomes.Offprint requests to: C. J. Bos     

Recombinant bacterial hemoglobin alters metabolism of Aspergillus niger

The filamentous fungus Aspergillus niger is used extensively for the production of enzymes and organic acids. A major problem in industrial fermentations with this fungus is to ensure sufficient supply of oxygen required for respiratory metabolism of the fungus. In case of oxygen limitation, the fungus will produce various by-products like organic acids and polyols. In order to circumvent this problem we here study the effects of the expression of a bacterial hemoglobin protein on the metabolism of A. niger. We integrated the vgb gene from Vitreoscilla sp. into the genome at the pyrA locus behind the strong gpdA promoter from Aspergillus nidulans. Analysis of secreted metabolites, oxygen uptake, CO(2) evolution and biomass formation points towards a relief of stress in the mutant expressing VHB when it is exposed to oxygen limitation. Our findings therefore point to an interesting strategy to attenuate unwanted side effects resulting from oxygen limitation during industrial fermentations with A. niger.     

Lipid metabolism in Aspergillus niger under conditions of heat shock

The processes of lipid synthesis and decomposition in Aspergillus niger under conditions of heat shock (HS) were studied in a pulse-chase experiment with ¹⁴C-labeled sodium acetate. HS (60 min) resulted in the synthesis of phospholipids and sphingolipids intensified compared to the control, as was evident from incorporation of the labeled substrate. The same pattern was observed for neutral lipids, especially for triacylglycerides, while incorporation of the label into sterols remained almost the same. Further cultivation for 3 h in the medium without the labeled substrate resulted in a significant decrease of the label content in the membrane lipids of both the control and the experiment, although under HS conditions this decrease was much more pronounced, especially for phosphatidylcholines and phosphatidylethanolamines. A threefold increase of the label content in phosphatidic acids was observed only under HS conditions. These results indicate more intense metabolism of the membrane lipids under heat shock and suggest the degradation of the major cell phospholipids as the factor responsible for the increased level of phosphatidic acids in A. niger mycelium.     

Reconstruction of the central carbon metabolism of Aspergillus niger.

The topology of central carbon metabolism of Aspergillus niger was identified and the metabolic network reconstructed, by integrating genomic, biochemical and physiological information available for this microorganism and other related fungi. The reconstructed network may serve as a valuable database for annotation of genes identified in future genome sequencing projects on aspergilli. Based on the metabolic reconstruction, a stoichiometric model was set up that includes 284 metabolites and 335 reactions, of which 268 represent biochemical conversions and 67 represent transport processes between the different intracellular compartments and between the cell and the extracellular medium. The stoichiometry of the metabolic reactions was used in combination with biosynthetic requirements for growth and pseudo-steady state mass balances over intracellular metabolites for the quantification of metabolic fluxes using metabolite balancing. This framework was employed to perform an in silico characterisation of the phenotypic behaviour of A. niger grown on different carbon sources. The effects on growth of single reaction deletions were assessed and essential biochemical reactions were identified for different carbon sources. Furthermore, application of the stoichiometric model for assessing the metabolic capabilities of A. niger to produce metabolites was evaluated by using succinate production as a case study.     

Rassenumwandlung bei Aspergillus niger

Zusammenfassung Es wurde versucht, zwei physiologisch durch die Oxalsäurebildung und die dadurch beeinflußte Richtung der Autolyse verschiedene Rassen von Aspergillus niger ineinander überzuführen. Dies gelang durch Einwirkung von Trocknen bzw. Erhitzen der Sporen und durch Passage über Tannin- und Oxalsäurelösung.Der Übergang der Rasse Göttingen neu (viel Oxalsäure, saure, helle Autolyse) in die Rasse Göttingen alt (wenig Oxalsäure, neutrale bzw. alkalische, dunkle Autolyse) ging bedeutend leichter als umgekehrt.Durch Oxalsäurepassage wird die Rasse Göttingen alt stärker in der Richtung der sauren, die Rasse Göttingen neu stärker in der Richtung der neutralen Autolyse beeinflußt.Die jeweiligen Veränderungen erwiesen sich drei bis acht Generationen, soweit untersucht, konstant.Mit Hilfe ähnlicher Methoden dürften die Veränderungen bei diesem Pilz der experimentellen Analyse zugänglich werden.Kurzer Auszug aus der Dissertation von Paul Hugo Kreutzmann, Untersuchungen über die Variabilität von Aspergillus niger. Göttingen 1935.     

Glucoamylase of Aspergillus niger

Aspergillus niger produces two extracellular glucoamylases (GAI of Mr 85 300 and GAII of Mr 77 600) separable on DEAE-cellulose. The enzymes differes in electrophoretic mobility, thermostability and substrate specificity. The GAI/GAII ratio depends on the concentration and form of nitrogen (nitrate or ammonium) in the culture medium. Proteinase VIII from Bacillus subtilis converts GAI to a form showing properties similar to those of GAII. Possible proteolytic degradation of GAI to GAII by Asp. niger endogenous proteinase(s) is suggested.     

Aspergillus niger sulfhydryl oxidase

A procedure for the isolation of a sulfhydryl oxidase from an Aspergillus niger cell suspension involved three major steps and yielded enzyme preparations exhibiting a single but diffuse protein-containing zone when subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with a subunit molecular weight estimated to be 53,000. Sedimentation equilibrium experiments indicated a native molecular weight of 106,000. Analyses for sugar residues showed that the enzyme is a glycoprotein, containing 20.3% neutral hexose and 1.9% aminohexose by weight. This enzyme catalyzed the conversion of reduced glutathione (GSH) to its disulfide form, with concomitant consumption of O2 and release of H2O2. The ratio of GSH consumed to H2O2 produced was determined to be 2:1. At 25 degrees C, the optimum pH for the oxidation of GSH was 5.5. Under these conditions, the enzyme had a Michaelis constant of 0.3 mM for GSH. Other low molecular weight thiol compounds (cysteine, dithiothreitol, and 2-mercaptoethanol) were also oxidized, but the Michaelis constants for these substrates were substantially higher than that for GSH under identical conditions of temperature and pH. The rate of reactivation of reductively denatured ribonuclease A was enhanced by the presence of sulfhydryl oxidase, indicating that the latter is capable of oxidizing protein-associated thiol groups. The UV-visible spectrum of sulfhydryl oxidase solution had absorbance maxima at 274, 364.5, and 442.5 nm and was otherwise characteristic of the spectra of known flavoproteins.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dye biosorption sites in Aspergillus niger

Aspergillus niger is capable of removing dyes from an aqueous solution. In the study, the roles played by three major functional groups: carboxyl, amino and phosphate, and the lipid fraction in the biomass of A. niger in biosorption of four dyes, Basic Blue 9, Acid Blue 29, Congo Red and Disperse Red 1, were investigated. These functional groups in A. niger were chemically modified individually to determine their contribution to the biosorption of dyes. It was found that biosorption of dyes was influenced by the functional groups in the fungal biomass and the chemical structure of the dyes.