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. 1978 Sep 15;174(3):1011–1019. doi: 10.1042/bj1741011

Sub-mitochondrial location of Ruthenium Red-sensitive calcium-ion transport and evidence for its enrichment in a specific population of rat liver mitochondria

Fyfe L Bygrave 1, Thomas P Heaney 1, Chidambaram Ramachandran 1
PMCID: PMC1186007  PMID: 728072

Abstract

1. Seven fractions sedimenting at between 3000 and 120000g-min were prepared from a rat liver homogenate by differential centrifugation in buffered iso-osmotic sucrose. The following measurements were carried out on each of these fractions: Ruthenium Red-sensitive Ca2+ transport in the absence and in the presence of Pi as well as in the presence of N-ethylmaleimide to prevent Pi cycling, succinate-supported respiration in the absence and in the presence of ADP, the ΔE and −59 ΔpH components of the protonmotive force, cytochrome oxidase, uncoupler-stimulated adenosine triphosphatase, α-glycerophosphate dehydrogenase, Pi content and the effect on the `resting' rate of respiration of repeated additions of a fixed Ca2+ concentration. 2. Ca2+ transport either in the presence or in the absence of added Pi and in the presence of N-ethylmaleimide exhibits significantly higher rates in the fraction sedimenting at 8000g-min. By contrast, respiration in the presence or in the absence of added ADP and the values for ΔE and −59 ΔpH were similar in those fractions sedimenting between 4000 and 20000g-min, indicating that the driving force for Ca2+ transport was similar in each of these fractions. 3. Experiments designed to determine the capacity of the individual fractions for Ca2+, as measured by the effect of repeated additions of Ca2+ on the resting rate of respiration, showed that fraction 2, i.e. that sedimenting at 8000g-min, also exhibited the greatest tolerance towards the uncoupling action of the ion. 4. Of the three enzyme activity profiles, only that of α-glycerophosphate dehydrogenase was similar to that of Ca2+ transport. Because previous workers have assigned this enzyme to loci in the inner peripheral membrane [Werner & Neupert (1972) Eur. J. Biochem. 25, 379–396], it is concluded that the Ruthenium Red-sensitive Ca2+- transport system also is located in this domain of the inner membrane. The relation of these findings to the mechanisms of mitochondrial Ca2+ transport and the biogenesis of mitochondria is discussed.

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Selected References

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