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.
D000596:Amino Acids / Q000032:analysis; D000937:Antigen-Antibody Reactions; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism; D008291:Malate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism; D008928:Mitochondria / Q000201:enzymology; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D000596:Amino Acids; D007527:Isoenzymes; D010446:Peptide Fragments; D008291:Malate Dehydrogenase