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Plant Physiology 51:1127-1132 (1973)
© 1973 American Society of Plant Biologists

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Articles

Malate Dehydrogenase Isoenzymes in Division Synchronized Cultures of Euglena

Barry Davis and M. J. Merrett

1 Postgraduate School of Studies in Biological Sciences, University of Bradford, Bradford, Yorkshire BD7 1DP, England

Sucrose density gradient centrifugation of broken cell suspensions of autotrophically grown Euglena gracilis Klebs. has allowed the separation of chloroplasts, mitochondria, and peroxisomes. Chlorophyll was taken as a marker for chloroplasts, fumarase and succinate dehydrogenase for mitochondria, and glycolate oxidoreductase for peroxisomes. Peaks of malate dehydrogenase (L-malate-NAD oxidoreductase, EC 1.1.1.37) activity were found in the mitochondrial and peroxisomal fractions. Acrylamide gel electrophoresis showed specific isoenzymes in the mitochondrial and peroxisomal fractions and a third isoenzyme in the supernatant. The mitochondrial isoenzyme which had a Km (oxaloacetate) of 30µM was inhibited by oxaloacetate concentrations above 0.17 mM, an inhibition of 50% being given by 0.9 mM oxaloacetate. The peroxisomal isoenzyme had a Km (oxaloacetate) of 24 µM, was inhibited by oxaloacetate concentrations above 0.13 mM, 50% inhibition being given by 0.25 mM oxaloacetate. Malate dehydrogenase activity in the supernatant did not show inhibition by increasing oxaloacetate concentration, the Km (oxaloacetate) being 91 µM.

In division synchronized cultures of Euglena, all three isoenzymes of malate dehydrogenase were synthesized over the light phase of the cycle. Darkening light phase cultures did not affect malate dehydrogenase activity. The addition to cultures of cycloheximide at a concentration previously shown to inhibit protein synthesis on Euglena cytoplasmic ribosomes completely inhibited increase in malate dehydrogenase activity over the cell cycle. Malate dehydrogenase activity was unaffected by the addition of chloramphenicol in amounts known to inhibit preferentially protein synthesis on 70S ribosomes.








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Copyright © 1973 by the American Society of Plant Biologists