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Advances in the Purification of the Mitochondrial Ca2+ Uniporter Using the Labeled Inhibitor 103Ru360

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Abstract

For many years the calcium uniporter has eluded attempts of purification, partly because of the difficulties inherent in the purification of low-abundance hydrophobic proteins (Reed and Bygrave, 1974). Liquid-phase preparative isoelectric focusing improved the fractionation of mitochondrial membrane proteins. A single 6-h run resulted in a 90-fold increase in specific activity of pooled active fractions over a semipurified fraction, allowing for enrichment of the calcium transport function in cytochrome oxidase vesicles. An additional powerful tool in the isolation of the uniporter was the use of the labeled inhibitor 103Ru360 as an affinity ligand; by following this procedure a protein of 18 kDa was purified in nondenatured, but rather inactive, form. The labeled protein corresponds to the protein that showed Ca2+ transport activity.

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Authors
  1. Cecilia Zazueta
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  2. Gabriela Zafra
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  3. Gabriela Vera
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  4. César Sánchez
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  5. Edmundo Chávez
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Zazueta, C., Zafra, G., Vera, G. et al. Advances in the Purification of the Mitochondrial Ca2+ Uniporter Using the Labeled Inhibitor 103Ru360 . J Bioenerg Biomembr 30, 489–498 (1998). https://doi.org/10.1023/A:1020546331217

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