Journal of Bioenergetics and Biomembranes

, Volume 26, Issue 2, pp 231–238

Purification of the channel component of the mitochondrial calcium uniporter and its reconstitution into planar lipid bilayers

  • Galina D. Mironova
  • Marc Baumann
  • Oleg Kolomytkin
  • Zoja Krasichkova
  • Adgemurat Berdimuratov
  • Tatjana Sirota
  • Ismo Virtanen
  • Nils-Erik L. Saris
Original Articles

DOI: 10.1007/BF00763072

Cite this article as:
Mironova, G.D., Baumann, M., Kolomytkin, O. et al. J Bioenerg Biomembr (1994) 26: 231. doi:10.1007/BF00763072

Abstract

The purification of the channel-forming component of the mitochondrial calcium uniporter and its channel properties are described. After ethanol and 50% ethanol-water extraction of mitochondria from beef heart or perfused rat liver, the extract was passed through thiopropyl-Sepharose 6B column, and absorbed components were eluted with 2-mercaptoethanol, followed by gel-filtration on Sephadex G-15. The last fraction eluted (Mr about 2000) was then subjected to reverse-phase high-performance liquid chromatography. Of the more than 10 distinct peaks, only one showed specific Ca2+-channel activity in BLM with properties similar to earlier, less extensively purified preparations, i.e., conductance of 20 pS and multiples thereof, clustering of channels, participation of 2 or more subunits in channel formation, and sensitivity to 1 µM ruthenium red. Voltage sensitivity and cooperativity between channels are described. The Ca2+-binding glycoprotein with which the peptide was associated was found to have high homology with human acid α1-glycoprotein (orosomucoid) and to show identity with beef plasma orosomucoid in the Ouchterlony immunodiffusion test.

Key words

Acid α1-glycoprotein black-lipid membrane calcium uniporter channel orosomucoid reconstitution 

Copyright information

© Plenum Publishing Corporation 1994

Authors and Affiliations

  • Galina D. Mironova
    • 1
  • Marc Baumann
    • 2
  • Oleg Kolomytkin
    • 3
  • Zoja Krasichkova
    • 1
  • Adgemurat Berdimuratov
    • 1
  • Tatjana Sirota
    • 1
  • Ismo Virtanen
    • 4
  • Nils-Erik L. Saris
    • 2
  1. 1.Institute of Theoretical and Experimental BiophysicsRussian Academy of SciencesPushchinoRussia
  2. 2.Department of Medical ChemistryUniversity of HelsinkiHelsinkiFinland
  3. 3.Institute of Cell BiophysicsRussian Academy of SciencesPushchinoRussia
  4. 4.Department of AnatomyUniversity of HelsinkiHelsinkiFinland