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D Mecke

Publications and source records attributed to D Mecke.

At least 73 records · Page 4Linked to original sources
PubMed1982

Genetic evidence for a role of hexokinase isozyme PII in carbon catabolite repression in Saccharomyces cerevisiae.

A mutant of Saccharomyces cerevisiae that was selected for resistance to carbon catabolite repression also had reduced hexokinase activity. Hexokinase isoenzymes were purified from mutant and wild type cells. The specific glucokinase and hexokinase isozyme PI were present at normal levels in mutant and wild type, but no hexokinase isozyme PII activity was detected in the mutant. Staining for enzyme activity after electrophoresis of crude extracts also indicated that hexokinase PII was absent in the mutant. Mutant and wild type segregants gained by tetrad analysis were investigated electrophoretically. Staining for enzyme activity confirmed that catalytically inactive hexokinase PII and the defect in carbon catabolite repression always co-segregated. The results support the hypothesis that hexokinase PII might mediate carbon catabolite repression.

Enzyme Repression↗
PubMed1982

The Triton X-100 and high salt resistant residue of Saccharomyces cerevisiae nuclear membranes.

Saccharomyces cerevisiae nuclear membranes were prepared from isolated nuclei by digesting chromatin with deoxyribonuclease after an initial treatment of nuclei with very diluted buffers. When the nuclear membranes were treated with 5% Triton X-100 and 1M NaCl an insoluble fibrous net was obtained which consisted mainly of protein with Mr values of 85 000, 48 000, 45 000, 39 000 and 31 000. Lamins, a set of proteins with Mr = 65 000--75 000, which were shown to be the major proteins of the insoluble nuclear membrane residue of higher eukaryotes, were not found.

Cell Fractionation↗
PubMed1982

Alterations in activity and ultrastructural localization of several phosphatases on the surface of adult rat hepatocytes in primary monolayer culture.

Several enzymes associated with the hepatocyte cell surface, alkaline phosphatase (AP), 5'-nucleotidase (5'N), Mg++- and total Na+K+Mg++-ATpase, were assayed and localized cytochemically in order to gain insight into alterations of the plasma membrane components during reassociation of hepatocytes in primary monolayer culture. During a period of 4 days the activities of 5'nucleotidase and alkaline phosphatase increased spontaneously up to three- and four-fold, respectively. Dexamethasone reinforce the rise of alkaline phosphatase activity but retarded the increase of that of 5'nucleotidase. However, after the third day the level of 5'nucleotidase activity converged with the untreated controls. The activities of Mg++- and Na+K+Mg++-ATPase, which closely paralleled each other, remained essentially unchanged throughout cultivation and were not affected by dexamethasone. Cytochemical demonstration of alkaline phosphatase, 5'nucleotidase and Mg++-ATPase, using the lead salt method, revealed the potential presence of reaction product on the whole cell surface. However, the cells did not react uniformly, particularly on bile canalicular membranes. This heterogeneity seems to be due to different stages of canalicular development and to different functional states of the cultured hepatocytes.

5'-Nucleotidase↗
PubMed1979

Activation of glutaminase by phosphoribosyl-pyrophosphate and its interference with the assay of phosphoribosylpyrophosphate amidotransferase.

Phosphate-dependent glutaminase (L-glutamine amidohydrolase, EC 3.5.1.2) from rat liver was found to be strongly activated by phosphoribosylpyrophosphate (P-rib-PP), the substrate of amidophosphoribosyltransferase (EC 2.4.2.14). Since the assay of the latter is based on the P-rib-PP-dependent conversion of glutamine to glutamate, the amidotransferase activities determined in crude tissue preparations were found to be too high. The interference of glutaminase, however, could be completely eliminated by its inactivation at 50 degrees C. Amidotransferase was not affected by the heat treatment. Because of the increased rate of the glutamate formation at this temperature, the incubation time of the assay could be significantly reduced.

Amidophosphoribosyltransferase↗
PubMed1979

Permissive effect of dexamethasone on glucagon induction of urea-cycle enzymes in perifused primary monolayer cultures of rat hepatocytes.

Parenchymal cells from adult rat liver, cultured in perifused monolayers, increased the levels of urea-cycle enzymes between 15% and 60% in response to glucagon within 24 h. This stimulation was drastically enhanced by the simultaneous presence of dexamethasone, especially in the case of argininosuccinate synthetase and argininosuccinate lyase, which increased nearly threefold. Dexamethasone itself produced only negligible stimulation, but exerted a similar effect on the stimulatory action of glucagon, if it was exclusively present during 6 h prior to the glucagon treatment, suggesting a permissive action of this hormone. The effect of glucagon, particularly in the presence of dexamethasone, was mimicked by dibutyryl adenosine 3':5'-monophosphate, whereas epinephrine was ineffective. All stimulations induced by hormones or dibutyryl adenosine 3':5'-monophosphate were abolished by cycloheximide, suggesting the involvement of protein synthesis in the induction process. Using the usual culture technique with a discontinuous supply of medium no significant effect of glucagon and dexamethasone could be measured. This striking difference between both culture systems indicates that perifusion is the more adequate in vitro system for studies of the regulation of enzyme levels. Possible reasons for the failure of hormonal stimulation of urea-cycle enzymes in normal monolayer culture are discussed.

Animals↗
PubMed1978

Evidence for catabolite degradation in the glucose-dependent inactivation of yeast cytoplasmic malate dehydrogenase.

The cytoplasmic malate dehydrogenase of Saccharomyces cerevisiae was radioactively labeled during its synthesis on a glucose-free derepression medium. After purification a sensitive radio-immunoassay for this enzyme could be developed. The assay showed that after the physiological, glucose-dependent 'catabolite inactivation' of cytoplasmic malate dehydrogenase an inactive enzyme protein is immunologically not detectable. Together with the irreversibility of this reaction in vivo this finding strongly suggest a proteolytic mechanism of enzyme inactivation. For this process the term 'catabolite degradation' is used.

Antibody Specificity↗
PubMed1978

The malate dehydrogenase isoenzymes of Saccharomyces cerevisiae. Purification, characterisation and studies on their regulation.

1. One mitochondrial and one cytoplasmic malate dehydrogenase isoenzyme could be purified from acetate grown cells of the yeast Saccharomyces cerevisiae. 2. The purification procedure uses chromatography on dextran blue columns as an essential step for enrichment, and reverse ammonium sulfate chromatography on celite for isoenzyme separation. 3. The homogeneity of the preparations was established by gel electrophoreses in the presence of sodium dodecylsulfate and by a sedimentation run in the analytical ultracentrifuge. 4. Both enzymes are dimers with a molecular weight of 75 000 for the cytoplasmic and of 68 000 for the mitochondrial enzyme. 5. Amino acid analysis and peptide mapping showed that both enzymes are closely related, but genetically different (true isoenzymes). 6. The cytoplasmic enzyme shows electrophoretic splitting. This is most likely due to post-translational deamination in vivo. 7. Antibodies to both isoenzymes could be obtained in rabbits. The antisera to cytoplasmic malate dehydrogenase were specific for this enzyme. Antisera to mitochondrial malate dehydrogenase react with both isoenzymes. Neither type of antisera precipitated an inactive protein after the glucose-dependent inactivation of cytoplasmic malate dehydrogenase in vivo.

Amino Acids↗
PubMed1977

In vivo and in vitro studies on the glucose dependent inactivation of yeast cytoplasmic malate dehydrogenase.

The cytoplasmic malate dehydrogenase in the yeast Saccharomyces cerevisiae is known to be inactivated by a glucose dependent process. In this paper it is shown that in vivo effectors of the glucose metabolism (arsenate, iodoacetate, acetaldhyde) inhibit the inactivation or change the inactivation kinetics. In vitro it was possible to inactivate the malate dehydrogenase by addition of the glucose metabolite glyceraldehyde 3-phosphate. The physiological relevance of this modification and the effect of malate dehydrogenase inactivation on the glyoxylate cycle in yeast is discussed.

Acetaldehyde↗
PubMed1977

Effect of insulin on glycogen and protein synthesis in monolayer cultures of hepatocytes from normal and alloxan diabetic rats.

The effects of insulin on net glycogen synthesis and amino acid incorporation into protein were studied in cultured hepatocytes from adult normal and alloxan diabetic rats. Insulin stimulated glycogen synthesis in monolayer cells throughout a four day culture period and enhanced leucine incorporation into protein more effectively in normal cells with high glycogen levels than in cultured diabetic cells. These differences correlate well with the observed cellular ultrastructures which were maintained much better in the presence of insulin. Restoration of the morphological changes of alloxan diabetic hepatocytes to normal liver cell structures can be observed at any time during the culture period by giving insulin continuously.

Animals↗