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J Visser

Publications and source records attributed to J Visser.

At least 199 records · Page 11Linked to original sources
PubMed1990

Purification and characterization of polygalacturonases produced by the hyphal fungus Aspergillus niger.

Five endo-polygalacturonases (poly(1,4-alpha-D-galacturonide) glycanohydrolase, EC 3.2.1.15) and one exo-polygalacturonase (poly(1,4-alpha-D-galacturonide) galacturonohydrolase, EC 3.2.1.67) were isolated from a commercial pectinase preparation derived from Aspergillus niger. All five endo-enzymes could be purified to homogeneity by affinity chromatography on cross-linked alginate, ion-exchange chromatography, chromatofocusing, and gel permeation chromatography. The exo-polygalacturonase was only partially purified but free from endo-polygalacturonase activity. The two most abundant endo-polygalacturonases (endo-I and endo-II), with molecular masses of 55 and 38 kDa, respectively, are quite different with respect to their isoelectric point, specific activity, mode of action on oligomeric substrates, and amino acid composition. The physicochemical properties of the other three endo-polygalacturonases (endo-IIIA, endo-IIIB, and endo-IV), present in low amounts, are quite similar to those of the endo-I type. The pH optima of all these endo-polygalacturonases are in the range of 4.3-4.9.

Alginates↗
PubMed1989

A study on the buffering activity of the human rectum. In vivo demonstration of HCO3- and H+ secretion after rectal application of fluids with an unphysiological pH.

The buffering activity of the human rectum was evaluated in various pilot studies, using a rectal lumen perfusion method. Quantitative alkaline and acid secretion was measured, respectively, by direct titration according to the pH-stat method. The change in composition of the perfusate with regard to the test drug benzoate and the concentration HCO3- was detected. The pilot studies suggest that rectal mucosa is capable of neutralizing a pH deviating from the physiological value within a remarkably short time. In the acid region, at least a part of the alkalizing activity is the result of HCO3- secretion. It has been shown that this process is stimulated by the H+ as well as Cl- ions. In the basic region neutralization very likely occurs by means of H+ secretion.

Bicarbonates↗
PubMed1989

Rhizosphere Acidification by Iron Deficient Bean Plants: The Role of Trace Amounts of Divalent Metal Ions: A Study on Roots of Intact Plants with the Use of C- and P-NMR.

Rhizosphere acidification by Fe-deficient bean (Phaseolus vulgaris L.) plants was induced by trace amounts of divalent metal ions (Zn, Mn). The induction of this Fe-efficiency reaction was studied by (14)CO(2) and (11)CO(2) fixation experiments, and with (31)P-NMR on roots of whole plants. The starting and ending of an acidification cycle was closely coupled to parallel changes in CO(2) fixation, within the maximal resolution capacity of 20 min. (31)P-NMR experiments on intact root systems showed one peak which was ascribed to vacuolar free phosphate. At the onset of proton extrusion this peak shifted, indicating increase of pH in the cells. Proton extrusion was inhibited, with a lag period of 2 hours, by the protein synthesis inhibitors cycloheximide and hygromycin. It is assumed that Zn and Mn induce proton extrusion in Fe-deficient bean roots by activating the synthesis of a short-living polypeptide; the NMR data suggest a role for this peptide in the functioning of a proton pumping ATPase in the plasma membrane.

Journal Article↗
PubMed1988

Isolation and transformation of the pyruvate kinase gene of Aspergillus nidulans.

The Aspergillus nidulans pyruvate kinase gene was isolated by heterologous hybridization using the corresponding yeast gene as a probe. A 2.9 kb EcoRI/BamHI fragment, which exclusively hybridized to the yeast gene, was subcloned in pBR322. This clone was used to transform an A. nidulans pkiA deletion mutant to PKI+. The analysis of transformants with respect to the kind of integration revealed about 80% homologous integration--55% by a double cross-over event (type III integration), 25% by a single cross-over event (type I integration). Type II transformants (20%) that arise by non-homologous integration have not been further characterized with respect to the sites of integration. A direct correlation between the number of copies of the gene integrated into the genome and the measured pyruvate kinase activity was found after growth ona glycolytic carbon source. From this, it was concluded that the 2.9 kb EcoRI/BamHI fragment contains the complete pyruvate kinase structural gene, including the promoter region. However, after growth on a gluconeogenic carbon source, the regulation of gene expression was found to be disturbed. On acetate an increase in activity per gene copy (0.2 IU) was found in the transformants, as compared with wild-type levels. It is suggested that the pyruvate kinase gene is regulated by negative control, and that some sequences involved in this regulation are missing in the cloned fragment.

Aspergillus nidulans↗
PubMed1988

Structure of the Aspergillus nidulans pyruvate kinase gene.

The complete nucleotide sequence of the Aspergillus nidulans pyruvate kinase gene, including its flanking sequences, is presented. The gene has a 1,578 bp coding sequence that encodes a protein of 526 amino acids; the latter is strongly homologous to the pyruvate kinases found in Saccharomyces cerevisiae (66%) and mammals (53%). The gene is interrupted by seven introns, three of which are in a conserved position compared to those present in the mammalian pyruvate kinase genes sequenced thus far. A fourth intron within the mononucleotide binding fold domain is in a conserved position with respect to the position of an intron within the NAD+ binding region of maize ADH I. The transcription start site has been determined; a major site of transcription was found 80 bp before the translation initiation codon. The promoter region of the A. nidulans pyruvate kinase gene contains no direct homologies with the TATA or CCAAT sequences in the expected region (30-70 bp) before the transcription initiation site. However, extended CT-enriched regions are found in the promotor region, similar to what has been observed in genes that are highly expressed in Saccharomyces cerevisiae and filamentous fungi.

Amino Acid Sequence↗
PubMed1988

Glycerol uptake mutants of the hyphal fungus Aspergillus nidulans.

A new class of glycerol non-utilizing mutants, designated glcC, has been isolated. The glcC gene was mapped in linkage group VI and mutants were found to complement the reference strains glcA1 (linkage group V) and glcB33 (linkage group I) in diploids. The new mutants were unable to grow on glycerol. However, in contrast to the glcA and glcB phenotype these mutants did grow well on dihydroxyacetone and D-galacturonate. By in vivo 13C NMR spectroscopy it was shown that the glcC mutant did not take up glycerol but did take up dihydroxyacetone. The latter substrate was converted intracellularly into glycerol which was then catabolized as normal.

Aspergillus nidulans↗
PubMed1988

A rapid purification procedure for pyruvate kinase from the hyphal fungus Aspergillus nidulans.

Pyruvate kinase was purified from the filamentous fungus Aspergillus nidulans with a 45-55% yield. The procedure involved dye-affinity chromatography and fast protein liquid chromatography, resulting in highly active and pure enzyme in milligram quantities within 2 days. The purified enzyme, a tetramer with a subunit molecular weight of 65,000 and an isoelectric point of 4.7, was used to determine the amino acid composition.

Amino Acids↗
PubMed1987

13C-NMR analysis of Aspergillus mutants disturbed in pyruvate metabolism.

The metabolic consequences of two defects in pyruvate metabolism of the hyphal fungus Aspergillus nidulans have been investigated by natural abundance 13C-NMR spectroscopy. A pyruvate dehydrogenase complex (pdh) mutant, grown on acetate, accumulates alanine upon starvation which is derived from mannitol reserves. The L-alanine level increases further upon incubation with the non-permissive substrate D-glucose. L-Glutamate is absent from these spectra as it is required both for the transamination of pyruvate and as a reaction on an impaired energy metabolism in such a pdh-deficient strain. A pyruvate carboxylase (pyc) mutant, grown upon acetate, only starts to accumulate alanine after a long incubation period with D-glucose, due to the long-lasting presence of phosphoenolpyruvate carboxykinase and malic enzyme, which are both induced by growth on acetate. When this strain is grown on D-fructose and L-glutamate, alanine also accumulates within 3 h upon transfer to D-glucose.

Acetates↗
PubMed1987

Determination of cyclodextrins in biological fluids by high-performance liquid chromatography with negative colorimetric detection using post-column complexation with phenolphthalein.

A rapid and sensitive high-performance liquid chromatographic method for the analysis of beta- and gamma-cyclodextrin in aqueous biological fluids such as plasma, urine, or tissue homogenate is described. The chromatographic system consists of a microBondapak Phenyl column as stationary phase and a mobile phase of water with 10% methanol. After post-column addition of an alkaline solution of phenolphthalein, negative colorimetric detection is used. The elution solvent and post-column reagent were mixed in a capillary tubing of 1.5 m (1.0 mm I.D.). Two methods of sample treatment are given, one for large (1.0 ml) and one for small (0.1 ml) sample volumes. Both methods were shown to be linear and reproducible. The detection limit for beta-cyclodextrin was 1.0 microgram/ml (0.77 nmol/ml). The method was used in the determination of some pharmacokinetic parameters of beta-cyclodextrin in rats after intravenous injection.

Animals↗
PubMed1986

Crossed affinity immunoelectrophoresis of the Escherichia coli pyruvate dehydrogenase complex.

The crossed immunoelectrophoretic pattern obtained with the intact pyruvate dehydrogenase complex of Escherichia coli can be modified when this technique is combined with affinity gel electrophoresis using reactive dyes coupled to agarose as ligands. The patterns that arise have been interpreted with respect to localization of the three component enzymes. This was realized by using antibodies with different specificity, active enzyme staining and E2-E3 subcomplex behaviour. Dissociation of E1 subunits occurs more easily than that of E3 but remains incomplete in this system. The free reactive Procion Blue-MX dyes tested inactivate the complex even at neutral pH. The dyes react with all three components but E3 (80%) and E2 (15-20%) retain part of their catalytic activity. Modification leads to an enhanced dissociation of E1.

Coloring Agents↗
PubMed1986

Characterization of Aspergillus nidulans mutants in carbon metabolism isolated after D-galacturonate enrichment.

A selective method for the isolation of Aspergillus nidulans mutants defective in the pyruvate dehydrogenase complex was devised. The essential steps in the procedure were a mutagenic treatment of conidia with X-rays to about 50% survival, followed by filtration enrichment in minimal medium with D-galacturonate as sole carbon source, and rescue on complete medium with acetate. The mutants thus isolated were phenotypically characterized on the basis of growth tests, and different genotypes were assigned on the basis of complementation tests. The majority of the mutants that were unable to utilize galacturonate were defective in one of the components of the pyruvate dehydrogenase complex. In addition, mutants defective in pyruvate carboxylase, mutants defective in glycerol catabolism and some novel mutants which were only unable to use D-galacturonate as carbon source were found. At least two genes were shown to be involved in D-galacturonate metabolism.

Aspergillus nidulans↗
PubMed1985

Gene amplification in Aspergillus nidulans by transformation with vectors containing the amdS gene.

Conidial protoplasts of an A. nidulans amdS deletion strain (MH1277) have been transformed to the AmdS+ phenotype with a plasmid carrying the wild type gene (p3SR2). Optimalisation of transformation and plating conditions now has resulted in frequencies of 300-400 transformants per microgram of DNA. Analysis of DNA from AmdS+ transformants of MH1277 showed that transformation had occurred by integration of vector DNA sequences into the genome. In virtually all these transformants multiple copies of the vector were present in a tandemly repeated fashion, not preferentially at the resident, partially deleted amdS gene. It is suggested that the observed integration phenomena are dependent on the genetic background of the A. nidulans strain, used for transformation. A model to explain the tandem type of integration is proposed.

Aspergillus nidulans↗
PubMed1985

Drastic improvement in the rectal absorption profile of morphine in man.

Rectal absorption of morphine HCl from aqueous vehicles at different pHs in man has been compared with an orally administered solution. Plasma concentrations of morphine were measured by electrochemical HPLC analysis after a single dose of 10 mg morphine HCl, in a cross-over study in 7 volunteers. Rectal absorption of morphine was dependent on pH, which could be explained as being due to pH partitioning. The absorption rate and bioavailability could be greatly improved, as compared to orally administered morphine, by adjusting the pH. It was concluded that a rectal solution adjusted to pH 7 to 8 provided an entirely adequate dosage form.

Administration, Oral↗
PubMed1985

Expression of an Escherichia coli beta-galactosidase fusion gene in Aspergillus nidulans.

We inserted in frame the Escherichia coli lacZ gene into the protein-coding region of the Aspergillus nidulans trpC gene and introduced the resultant fused gene into the A. nidulans genome. A functional beta Gal fusion protein was produced. Removal of the trpC transcription and translation initiation sequences from the fusion gene abolished production of the fusion protein, showing that expression is dependent on these sequences. Thus, lacZ fusions should be of use for estimating gene activity in a. nidulans.

Aspergillus nidulans↗
PubMed1985

13C NMR studies of carbon metabolism in the hyphal fungus Aspergillus nidulans.

Natural-abundance high-resolution 13C NMR spectra (linewidth, 10 Hz) of the hyphal fungus Aspergillus nidulans have been obtained after growth on glycolytic or gluconeogenic carbon sources. Various polyols, some tricarboxylic acid-cycle intermediates and amino acids, and some phospholipids and fatty acyl compounds are present. The polyols found are mannitol, arabitol, erythritol, and glycerol. The nature of the carbon source has a pronounced effect on the pool sizes of the various polyols. All are present when the fungus is grown on sucrose or sucrose/acetate under strongly aerobic conditions. When grown on acetate, both arabitol and glycerol levels are low, whereas on glycerol erythritol is also hardly detectable. The effect of oxygen is most outspoken in glycolytically grown cultures. Limited oxygenation leads to low levels of arabitol and glycerol. Strong oxygenation changes the ratio of erythritol to mannitol, favoring the C4 polyol. An increase in oxygen supply leads to (i) stimulation of the fluxes through the pentose phosphate pathway and glycolysis, (ii) an overflow of reduced metabolites both at the pentose phosphate pathway level (erythritol and arabitol) and at the C3 level of the glycolytic pathway (glycerol), and (iii) the usual accumulation of mannitol. Upon starvation, glycerol, the other three polyols, and the tricarboxylic acid-cycle intermediates and their associated amino acids disappear in this order. As in yeast, gluconeogenic substrates lead to the synthesis of trehalose, which also occurs when mycelium is grown on acetate/sucrose under limiting aeration. A transient formation of trehalose has been observed upon incubation of starved mycelium, cultured on different substrates, with [13C]glucose.

Aerobiosis↗