Summary
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1.
In the colourless acetate flagellate Polytomella caeca (Volvocales) 70–90% of the catalase activity and 60–90% of the uricase activity are particulate. After centrifugation of a crude particulate fraction on a sucrose gradient for 300 min at 60,000xg the distribution patterns of these microbodial marker enzymes overlap extensively those of cytochrome oxidase and succinic dehydrogenase marking the mitochondria. However, by centrifugation at 30,000 x g for 60 min a clear separation of the activity peaks of catalase and uricase from those of the mitochondrial enzymes can be achieved proving the existence of microbodies in Polymella caeca. After centrifugation at 60,000 x g for 15 hrs the microbodies were found around a density of 1.225 g/cm3.
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2.
On an average more than 90% of the activity of the glyoxylate cycle enzymes isocitrate lyase and malate synthetase are found in the soluble fraction. But after washing the particulate fraction twice the specific activity of isocitrate lyase in this fraction drops only by half. After centrifugation (60,000 x g, 300 min) of a particulate fraction on a sucrose gradient a small peak of isocitrate lyase activity can be observed below the mitochondria fraction around a density of 1.24 g/cm3 where no peak of catalase or uricase activity occurs. Therefore, the small amount of particulate isocitrate lyase is not associated with the microbodies.
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3.
30–40% of the acetyl-CoA synthetase are recovered in the particulate fraction. On sucrose gradients this enzyme shows distribution patterns very similar to cytochrome oxidase demonstrating its localization in the mitochondria. No peak or shoulder of the activity occurs in any other region of the gradient (except the top fraction) although an extramitochondrial activity of this enzyme has to be assumed.
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4.
The experiments indicate that in Polytomella caeca the microbodies do not have a direct function in the metabolism of acetate.
Zusammenfassung
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1.
In dem farblosen Acetatflagellaten Polytomella caeca (Volvocales) wurde unter Zugrundelegung von Katalase und Uricase als Leitenzymen das Vorkommen von Microbodies nachgewiesen. Nach Zentrifugation an einem Saccharose-Gradienten bei 60 000 x g für 15 Std sedimentieren die Microbodies bei einer Dichte von 1.225 g/cm3.
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2.
Über 90% der Aktivitäten der Schlüsselenzyme des Glyoxylatcyclus wurden in der löslichen Fraktion eines Homogenats gefunden. Die geringe Aktivität der Isocitratlyase in der partikulären Fraktion wrwies sich als relativ fest gebunden, und nach zweimaligem Waschen der Partikelfraktion nahm die spezifische Aktivität nur um die Hälfte ab. An Saccharose-Gradienten trat nach Zentrifugation bei 60 000 x g für 300 min ein Gipfel der Isocitratlyaseaktivität bei der Dichte 1.24 g/cm3 auf.
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3.
Partikuläre Acetyl-CoA-Synthetase konnte nur in den Mitochondrien lokalisiert werden.
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4.
Für Polytomella caeca konnte eine unmittelbare Funktion der Microbodies im Acetatstoffwechsel nicht nachgewiesen werden.
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Gerhardt, B. Zur Lokalisation von Enzymen der Microbodies in Polytomella caeca . Archiv. Mikrobiol. 80, 205–218 (1971). https://doi.org/10.1007/BF00410122
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DOI: https://doi.org/10.1007/BF00410122