Isoenzymes of Malate Dehydrogenase in <i>Saccharomyces cerevisiae</i>

Atzpodien, W.; Gancedo, Juana M.; Duntze, W.; Holzer, Helmut

doi:10.1111/j.1432-1033.1968.tb19573.x

Cited by 43 publications

(19 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, a reproducible method for quantitative isoenzyme separation from crude extracts was lacking. This increased the risk that an isoenzyme would be overlooked which is present in minor quantities [5].…”

Section: Purijicutinnmentioning

confidence: 99%

“…Witt et al [2] could show that glucosergrown cells contain only mitochondrial enzyme activity, whereas cells from a glucose-free medium contain malate dehydrogenase activity in the mitochondrial and in the cytoplasmic fraction. It could not definitely be shown whether the cytoplasmic activity stems from one or several isoenzymes [5,8]. Addition of a fermentable sugar to acetate-grown cells caused repression and a rapid drop of the cytoplasmic malate dehydrogenase activity [I, 3,4].…”

mentioning

confidence: 99%

“…Both enzymes are dimers with a molecular weight of 75000 for the cytoplasmic and of 68000 for the mitochondrial enzyme. 5. Amino acid analysis and peptide mapping showed that both enzymes are closely related, but genetically different (true isoenzymes).…”

mentioning

confidence: 99%

See 2 more Smart Citations

The Malate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae. Purification, Characterisation and Studies on Their Regulation

1978

View full text Add to dashboard Cite

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.

show abstract

Section: Purijicutinnmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

The Malate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae. Purification, Characterisation and Studies on Their Regulation

1978

View full text Add to dashboard Cite

show abstract

“…1). One branch is oxidative, leading to 2-OG formation (Nunez de Castro et al, 1970), whereas the other is reductive, leading to fumarate formation (Atzpodien et al, 1968;Sols et al, 1971). This was confirmed by analysing the metabolic network in S. cerevisiae during respiro-fermentative growth on glucose (Gombert et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

Investigation by 13C-NMR and tricarboxylic acid (TCA) deletion mutant analysis of pathways for succinate formation in Saccharomyces cerevisiae during anaerobic fermentation

2003

View full text Add to dashboard Cite

NMR isotopic filiation of 13 C-labelled aspartate and glutamate was used to explore the tricarboxylic acid (TCA) pathway in Saccharomyces cerevisiae during anaerobic glucose fermentation. The assimilation of [3-13 C]aspartate led to the formation of [2,3-13 C]malate and [2,3-13 C]succinate, with equal levels of 13 C incorporation, whereas site-specific enrichment on C-2 and C-3 of succinate was detected only with [3-13 C]glutamate. The non-random distribution of 13 C labelling in malate and succinate demonstrates that the TCA pathway operates during yeast fermentation as both an oxidative and a reductive branch. The observed 13 C distribution suggests that the succinate dehydrogenase (SDH) complex is not active during glucose fermentation. This hypothesis was tested by deleting the SDH1 gene encoding the flavoprotein subunit of the SDH complex. The growth, fermentation rate and metabolite profile of the sdh1 mutant were similar to those of the parental strain, demonstrating that SDH was indeed not active. Filiation experiments indicated the reductive branch of the TCA pathway was the main pathway for succinate production if aspartate was used as the nitrogen source, and that a surplus of succinate was produced by oxidative decarboxylation of 2-oxoglutarate if glutamate was the sole nitrogen source. Consistent with this finding, a kgd1 mutant displayed lower levels of succinate production on glutamate than on other nitrogen sources, and higher levels of oxoglutarate dehydrogenase activity were observed on glutamate. Thus, the reductive branch generating succinate via fumarate reductase operates independently of the nitrogen source. This pathway is the main source of succinate during fermentation, unless glutamate is the sole nitrogen source, in which case the oxidative decarboxylation of 2-oxoglutarate generates additional succinate.

show abstract

“…S. cerevisiae can utilize acetate and ethanol as carbon sources by inducing synthesis of isocitrate lyase and malate synthase (30), key glyoxylate cycle enzymes not expressed in mammalian cells, but whether the requirement for malate dehydrogenase activity in this cycle is fulfilled by the cytosolic isozyme or by a unique glyoxysomal enzyme has not been established. Various analyses of the isozymes of malate dehydrogenase in S. cerevisiae have been reported, with contradictory conclusions that this yeast contains two (21,30) or three (1) biochemically distinct enzymes. To resolve this question and to compare the structures and expression of the yeast enzymes, we have purified the major nonmitochondrial isozyme from yeast cells grown on acetate and have cloned the corresponding gene.…”

mentioning

confidence: 99%

Isolation, nucleotide sequence analysis, and disruption of the MDH2 gene from Saccharomyces cerevisiae: evidence for three isozymes of yeast malate dehydrogenase.

Minard¹,

McAlister-Henn²

1991

Mol. Cell. Biol.

View full text Add to dashboard Cite

The major nonmitochondrial isozyme of malate dehydrogenase (MDH2) in Saccharomyces cerevisiae cells grown with acetate as a carbon source was purified and shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to have a subunit molecular weight of approximately 42,000. Enzyme assays and an antiserum prepared against the purified protein were used to screen a collection of acetate-nonutilizing (acetate-) yeast mutants, resulting in identification of mutants in one complementation group that lack active or immunoreactive MDH2. Transformation and complementation of the acetate-growth phenotype was used to isolate a plasmid carrying the MDH2 gene from a yeast genomic DNA library. The amino acid sequence derived from complete nucleotide sequence analysis of the isolated gene was found to be extremely similar (49% residue identity) to that of yeast mitochondrial malate dehydrogenase (molecular weight, 33,500) despite the difference in sizes of the two proteins. Disruption of the MDH2 gene in a haploid yeast strain produced a mutant unable to grow on minimal medium with acetate or ethanol as a carbon source. Disruption of the MDH2 gene in a haploid strain also containing a disruption in the chromosomal MDHI gene encoding the mitochondrial isozyme produced a strain unable to grow with acetate but capable of growth on rich medium with glycerol as a carbon source. The detection of residual malate dehydrogenase activity in the latter strain confirmed the existence of at least three isozymes in yeast cells.Two isozymes of malate dehydrogenase in mammalian cells catalyze the interconversion of malate plus NAD+ and oxaloacetate plus NADH. The mitochondrial isozyme functions in the tricarboxylic acid cycle, and the cytosolic isozyme catalyzes a step in gluconeogenesis from pyruvate. Also, as components of the malate/aspartate shuttle cycle, the malate dehydrogenase isozymes control the exchange of metabolic intermediates and reducing equivalents across the mitochondrial membrane. Analyses of amino acid sequences for the isozymes from porcine heart (4, 5) and of nucleotide sequences for the porcine and murine cDNAs (25,26) (MATo leu2 his3-AJ trpl gcrl-J-J; 34) was used for isolation of the nonmitochondrial isozyme of malate dehydrogenase. Yeast strains S173-6B (MATa leu2-3,112 his3-AJ ura3-57 trpl-289; 7) and SAMDH1, a derivative of S173-6B containing a chromosomal disruption of the MDH1 gene (47), were used for transformation, growth studies, and cell fractionation. The parental S. cerevisiae strain MMY011 (MATa ade2-1 his3-11,15 leu2-3,112 trpl-l ura3-1 canl-100) and 370 on April 28, 2019 by guest

show abstract

Isoenzymes of Malate Dehydrogenase in Saccharomyces cerevisiae

Cited by 43 publications

References 22 publications

The Malate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae. Purification, Characterisation and Studies on Their Regulation

The Malate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae. Purification, Characterisation and Studies on Their Regulation

Investigation by 13C-NMR and tricarboxylic acid (TCA) deletion mutant analysis of pathways for succinate formation in Saccharomyces cerevisiae during anaerobic fermentation

Isolation, nucleotide sequence analysis, and disruption of the MDH2 gene from Saccharomyces cerevisiae: evidence for three isozymes of yeast malate dehydrogenase.

Contact Info

Product

Resources

About