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Trapping Oxidative Folding Intermediates During Translocation to the Intermembrane Space of Mitochondria: In Vivo and In Vitro Studies

  • Dionisia P. Sideris
  • Kostas Tokatlidis
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 619)

Abstract

The MIA40 pathway is a novel import pathway in mitochondria specific for cysteine-rich proteins of the intermembrane space (IMS). The newly synthesised precursors are trapped in the IMS by a disulfide relay mechanism that involves introduction of disulfides from the sulfhydryl oxidase Erv1 to the redox-regulated import receptor Mia40 and then on to the substrate. This thiol–disulfide exchange mechanism is essential for the import and oxidative folding of the incoming cysteine-rich substrate proteins. In this chapter we will describe the experimental methods that have been developed in order to study and characterise disulfide-trapped intermediates in yeast mitochondria.

Key words

Mitochondrial protein import mitochondrial intermembrane space MIA40 pathway Blue Native PAGE thiol-disulfide exchange mixed disulfide intermediates 

Notes

Acknowledgments

This work was supported by intramural funds from IMBB-FORTH, the University of Crete and the European Social Fund and national resources (to KT). DS was supported by a PENED grant. We are grateful to members of our lab for comments.

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Dionisia P. Sideris
    • 1
  • Kostas Tokatlidis
    • 2
  1. 1.Department of BiologyUniversity of Crete and Institute of Molecular Biology and Biotechnology-Foundation for Research and Technology HellasHeraklionGreece
  2. 2.Department of Materials Science and TechnologyUniversity of Crete and Institute of Molecular Biology and Biotechnology-Foundation for Research and Technology HellasHeraklionGreece

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