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Garedew 2006 Biochim Biophys Acta

From Bioblast
Publications in the MiPMap
Garedew A, Lemieux H, Schachner T, Blier PU, Tardif J-C, Gnaiger E (2006) High excess capacity of cytochrome c oxidase in permeabilized fibers of the mouse heart. Biochim. Biophys. Acta, EBEC Short Reports Suppl. 14 (2006): 167-168.

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Garedew A, Lemieux H, Schachner T, Blier PU, Tardif J-C, Gnaiger E (2006) Biochim. Biophys. Acta

Abstract: Metabolic flux control analysis and the concept of excess capacity of enzymes over pathway flux are related by the functional threshold, at which damage or inhibition of an enzyme reduces excess capacity to a minimum and starts to limit overall flux through the pathway. Excess capacity of cytochrome c oxidase (COX) varies between tissues, but little is known about differences between species. In particular, information is lacking on mitochondrial respiratory function in the mouse heart, despite the fact that transgenic mice provide increasingly important animal models. Permeabilized muscle fibers were prepared from the left ventricle of a single mouse heart, and measured in OROBOROS Oxygraph-2k instruments in parallel at 4, 25, 30, 37 and 40 °C (N³4). Threshold plots were constructed from azide titrations of flux through the electron transport chain (parallel e-input into complexes I+II with malate+pyruvate+glutamate+succinate and uncoupling by FCCP), versus COX (0.5 mM TMPD+2 mM ascorbate after uncoupling and inhibition by rotenone+malonate+antimycin A). Azide was used, since inhibition of COX by cyanide is reversed by pyruvate particularly at low oxygen levels. The inhibition constant, Ki, of COX for azide was 0.1 mM at 37 °C, increasing from 4 to 40 °C over two orders of magnitude. COX velocity measured with TMPD+ascorbate was 1.3-fold of maximum electron transport capacity of the respiratory chain at 25 to 40 °C, and 3.3-fold at 4 °C. In contrast, linear extrapolations of threshold plots revealed a COX excess capacity of 1.6-fold over pathway flux in the range of 30 to 40 °C, increasing to 1.8- and 7.6-fold at 25 °C and 4 °C, respectively. Application of complex I substrates only, would yield an apparent COX excess capacity of >3-fold over pathway flux (at 30 and 37 °C), since parallel e-input through complex I+II doubled flux compared to complex I substrates. Taken together, COX excess capacity in myocardial fibers of the mouse was significantly higher than in fibers of rat heart or human skeletal muscle. Results obtained under hypothermic incubation conditions of permeabilzed fibers may be extrapolated to physiological temperature of 37 °C with caution only. The very high COX excess capacity under hypothermia (4 °C) may compensate for hypothermic hypoxia by decreasing the p50 of mitochondrial respiration in parallel to the decreased p50 of hemoglobin and myoglobin. The present study yields an important baseline for further investigations of mitochondrial function in the mouse heart, including genetic models of acquired and inherited mitochondrial defects.


O2k-Network Lab: AT_Innsbruck_Gnaiger E, CA_Rimouski_Blier PU, CA_Edmonton_Lemieux H


Labels:


Organism: Mouse  Tissue;cell: Cardiac Muscle"Cardiac Muscle" is not in the list (Heart, Skeletal muscle, Nervous system, Liver, Kidney, Lung;gill, Islet cell;pancreas;thymus, Endothelial;epithelial;mesothelial cell, Blood cells, Fat, ...) of allowed values for the "Tissue and cell" property.  Preparation: Permeabilized Cell or Tissue; Homogenate"Permeabilized Cell or Tissue; Homogenate" is not in the list (Intact organism, Intact organ, Permeabilized cells, Permeabilized tissue, Homogenate, Isolated mitochondria, SMP, Chloroplasts, Enzyme, Oxidase;biochemical oxidation, ...) of allowed values for the "Preparation" property., Oxidase; Biochemical Oxidation"Oxidase; Biochemical Oxidation" is not in the list (Intact organism, Intact organ, Permeabilized cells, Permeabilized tissue, Homogenate, Isolated mitochondria, SMP, Chloroplasts, Enzyme, Oxidase;biochemical oxidation, ...) of allowed values for the "Preparation" property., Enzyme 

Regulation: Respiration; OXPHOS; ETS Capacity"Respiration; OXPHOS; ETS Capacity" is not in the list (Aerobic glycolysis, ADP, ATP, ATP production, AMP, Calcium, Coupling efficiency;uncoupling, Cyt c, Flux control, Inhibitor, ...) of allowed values for the "Respiration and regulation" property., Flux Control; Additivity; Threshold; Excess Capacity"Flux Control; Additivity; Threshold; Excess Capacity" is not in the list (Aerobic glycolysis, ADP, ATP, ATP production, AMP, Calcium, Coupling efficiency;uncoupling, Cyt c, Flux control, Inhibitor, ...) of allowed values for the "Respiration and regulation" property. 


HRR: Oxygraph-2k