Regulation of citric acid cycle by calcium

B. Wan, K. F. LaNoue, J. Y. Cheung, R. C. Scaduto

Research output: Contribution to journalArticle

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Abstract

The relationship of extramitochondrial Ca2+ to intramitochondrial Ca2+ and the influence of intramitochondrial free Ca2+ concentrations on various steps of the citric acid cycle were evaluated. Ca2+ was measured using the Ca2+ sensitive fluorescent dye fura-2 trapped inside the rat heart mitochondria. The rate of utilization of specific substrates and the rate of accumulation of citric acid cycle intermediates were measured at matrix free Ca2+ ranging from 0 to 1.2 μM. A change in matrix free Ca2+ from 0 to 0.3 μM caused a 135% increase in ADP stimulated oxidation of 0.6 mM α-ketoglutarate (K0.5 = 0.15 μM). In the absence of ADP and the presence of 0.6 mM α-ketoglutarate, Ca2+ (0.3 μM) increased NAD(H) reduction from 0 to 40%. On the other hand, when pyruvate (10 μM to 5 mM) was substrate, pyruvate dehydrogenase flux was insensitive to Ca2+ and isocitrate dehydrogenase was sensitive to Ca2+ only in the presence of added ADP. In separate experiments pyruvate dehydrogenase activation (dephosphorylation) was measured. Under the conditions of the present study, pyruvate dehydrogenase was found to be almost 100% activated at all levels of Ca2+, thus explaining the Ca2+ insensitivity of the flux measurements. However, if the mitochondria were incubated in the absence of pyruvate, with excess α-ketoglutarate and excess ATP, the pyruvate dehydrogenase complex was only 20% active in the absence of added Ca2+ and activity increased to 100% at 2 μM Ca2+. Activation by Ca2+ required more Ca2+ (K0.5 = 1 μM) than for α-ketoglutarate dehydrogenase. The data suggest that in heart mitochondria α-ketoglutarate dehydrogenase may be a more physiologically relevant target of Ca2+ action than pyruvate dehydrogenase.

Original languageEnglish (US)
Pages (from-to)13430-13439
Number of pages10
JournalJournal of Biological Chemistry
Volume264
Issue number23
StatePublished - Jan 1 1989

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Citric Acid Cycle
Pyruvic Acid
Oxidoreductases
Calcium
Mitochondria
Adenosine Diphosphate
Heart Mitochondria
Chemical activation
Fluxes
Pyruvate Dehydrogenase Complex
Isocitrate Dehydrogenase
Fura-2
Substrates
Fluorescent Dyes
NAD
Rats
Adenosine Triphosphate
Oxidation
Experiments

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Wan, B., LaNoue, K. F., Cheung, J. Y., & Scaduto, R. C. (1989). Regulation of citric acid cycle by calcium. Journal of Biological Chemistry, 264(23), 13430-13439.
Wan, B. ; LaNoue, K. F. ; Cheung, J. Y. ; Scaduto, R. C. / Regulation of citric acid cycle by calcium. In: Journal of Biological Chemistry. 1989 ; Vol. 264, No. 23. pp. 13430-13439.
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abstract = "The relationship of extramitochondrial Ca2+ to intramitochondrial Ca2+ and the influence of intramitochondrial free Ca2+ concentrations on various steps of the citric acid cycle were evaluated. Ca2+ was measured using the Ca2+ sensitive fluorescent dye fura-2 trapped inside the rat heart mitochondria. The rate of utilization of specific substrates and the rate of accumulation of citric acid cycle intermediates were measured at matrix free Ca2+ ranging from 0 to 1.2 μM. A change in matrix free Ca2+ from 0 to 0.3 μM caused a 135{\%} increase in ADP stimulated oxidation of 0.6 mM α-ketoglutarate (K0.5 = 0.15 μM). In the absence of ADP and the presence of 0.6 mM α-ketoglutarate, Ca2+ (0.3 μM) increased NAD(H) reduction from 0 to 40{\%}. On the other hand, when pyruvate (10 μM to 5 mM) was substrate, pyruvate dehydrogenase flux was insensitive to Ca2+ and isocitrate dehydrogenase was sensitive to Ca2+ only in the presence of added ADP. In separate experiments pyruvate dehydrogenase activation (dephosphorylation) was measured. Under the conditions of the present study, pyruvate dehydrogenase was found to be almost 100{\%} activated at all levels of Ca2+, thus explaining the Ca2+ insensitivity of the flux measurements. However, if the mitochondria were incubated in the absence of pyruvate, with excess α-ketoglutarate and excess ATP, the pyruvate dehydrogenase complex was only 20{\%} active in the absence of added Ca2+ and activity increased to 100{\%} at 2 μM Ca2+. Activation by Ca2+ required more Ca2+ (K0.5 = 1 μM) than for α-ketoglutarate dehydrogenase. The data suggest that in heart mitochondria α-ketoglutarate dehydrogenase may be a more physiologically relevant target of Ca2+ action than pyruvate dehydrogenase.",
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Wan, B, LaNoue, KF, Cheung, JY & Scaduto, RC 1989, 'Regulation of citric acid cycle by calcium', Journal of Biological Chemistry, vol. 264, no. 23, pp. 13430-13439.

Regulation of citric acid cycle by calcium. / Wan, B.; LaNoue, K. F.; Cheung, J. Y.; Scaduto, R. C.

In: Journal of Biological Chemistry, Vol. 264, No. 23, 01.01.1989, p. 13430-13439.

Research output: Contribution to journalArticle

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Wan B, LaNoue KF, Cheung JY, Scaduto RC. Regulation of citric acid cycle by calcium. Journal of Biological Chemistry. 1989 Jan 1;264(23):13430-13439.