Proteomics of Mitochondria
EMSL Project ID
9102
Abstract
The mitochondrial organelle originated when aerobic bacteria colonized primordial anaerobic eukaryotic cells, and then evolved to effectively generate cellular energy via aerobic glycolysis. Mitochondrial core functions such as oxidative phosphorylation, amino acid metabolism, fatty acid oxidation, and iron-sulfur cluster assembly are highly conserved from Saccharomyces cerevisiae to Homo sapiens. Therefore, missing components of the human mitochondrion can be identified in model organisms, such as yeast, and help defining their role in mitochondrial diseases. Our collaborative work with Pacific Northwest National Laboratory identified 546 mitochondrial proteins in yeast using reversed phase high resolution liquid chromatography (LC)/tandem mass spectrometry (MS/MS) and LC/Fourier Transform-Ion Cyclotron Resonance mass spectrometry (FTICR) applied on purified mitochondrial organelles. An integrative analysis of the proteins identified by MS with other systematic studies helped to define the parts list of the mitochondrial proteome in yeast (~700 proteins). We propose to continue our collaboration with PNNL and further identify mitochondrial proteins and study their function in various species. In the first part of the project we are interested to improve the coverage of AMT?s (Accurate Mass and Time Tags) for mitochondrial protein in yeast. In addition, the generation for AMT?s of mitochondrial protein in mouse is proposed. To enable these measurements, we will provide PNNL with highly purified mitochondrial organelles from yeast and mouse, respectively. These studies are prerequisite for future experiments on the quantification of mitochondrial proteins under various conditions and in different species. We would like to pilot such a quantitative analysis on a well known biological system: the mitochondrial TOM complex in Neurospora crassa which is conserved in eukaryotes. We will provide PNNL with highly purified TOM complex and mitochondrial outer membrane vesicles (OMV?s) containing the TOM complex. Because the identity and stoichiometry of the TOM subunits is known, quantitative measurements of the purified complex and OMV proteins in comparison will help to evaluate MS proteomics and current protein labeling technologies. These findings will help to implement proteomics technologies to more complex biological systems including the study of the human mitochondrial proteome. Please see the attached file for further details.
Project Details
Project type
Exploratory Research
Start Date
2004-05-21
End Date
2007-05-27
Status
Closed
Released Data Link
Team
Principal Investigator