The Effect of Calcium Ions on the Activity of the Pyruvate Dehydrogenase Complex in Ehrlich Ascites Tumor Cells

LP Dolgacheva, BN Goldstein, II Goryanin, VP Zinchenko

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

The metal-ion requirement of extracted and partially purified pyruvate dehydrogenase phosphate phosphatase from rat epididymal fat-pads was investigated with pig heart pyruvate dehydrogenase [32P]phosphate as substrate. The enzyme required Mg2+ (Km 0.5mm) and was activated additionally by Ca2+ (Km 1μm) or Sr2+ and inhibited by Ni2+. Isolated fat-cell mitochondria, like liver mitochondria, possess a respiration- or ATP-linked Ca2+-uptake system which is inhibited by Ruthenium Red, by uncouplers when linked to respiration, and by oligomycin when linked to ATP. Depletion of fat-cell mitochondria of 75% of their total magnesium content and of 94% of their total calcium content by incubation with the bivalent-metal ionophore A23187 leads to complete loss of pyruvate dehydrogenase phosphate phosphatase activity. Restoration of full activity required addition of both MgCl2 and CaCl2. SrCl2 could replace CaCl2 (but not MgCl2) and NiCl2 was inhibitory. The metal-ion requirement of the phosphatase within mitochondria was thus equivalent to that of the extracted enzyme. Insulin activation of pyruvate dehydrogenase in rat epididymal fat-pads was not accompanied by any measurable increase in the activity of the phosphatase in extracts of the tissue when either endogenous substrate or 32P-labelled pig heart substrate was used for assay. The activation of pyruvate dehydrogenase in fat-pads by insulin was inhibited by Ruthenium Red (which may inhibit cell and mitochondrial uptake of Ca2+) and by MnCl2 and NiCl2 (which may inhibit cell uptake of Ca2+). It is concluded that Mg2+ and Ca2+ are cofactors for pyruvate dehydrogenase phosphate phosphatase and that an increased mitochondrial uptake of Ca2+ might contribute to the activation of pyruvate dehydrogenase by insulin.
Original languageEnglish
Pages (from-to)1063-1067
Number of pages5
JournalBiochemistry (Moscow)
Volume61
Issue number8
Publication statusPublished - 1996

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