Mitochondria consist of four compartments-outer membrane, intermembrane space, inner membrane, and matrix-with crucial but distinct functions for numerous cellular processes. A comprehensive characterization of the proteome of an individual mitochondrial compartment has not been reported so far. We used a eukaryotic model organism, the yeast Saccharomyces cerevisiae, to determine the proteome of highly purified mitochondrial outer membranes. We obtained a coverage of ϳ85% based on the known outer membrane proteins. The proteome represents a rich source for the analysis of new functions of the outer membrane, including the yeast homologue (Hfd1/Ymr110c) of the human protein causing Sjö gren-Larsson syndrome. Surprisingly, a subclass of proteins known to reside in internal mitochondrial compartments were found in the outer membrane proteome. These seemingly mislocalized proteins included most top scorers of a recent genome-wide analysis for mRNAs that were targeted to mitochondria and coded for proteins of prokaryotic origin. Together with the enrichment of the precursor form of a matrix protein in the outer membrane, we conclude that the mitochondrial outer membrane not only contains resident proteins but also accumulates a conserved subclass of preproteins destined for internal mitochondrial compartments.

Background: Mass spectrometry based quantification of peptides can be performed using the iTRAQ™ reagent in conjunction with mass spectrometry. This technology yields information about the relative abundance of single peptides. A method for the calculation of reliable quantification information is required in order to obtain biologically relevant data at the protein expression level.

To study human cancer development, cell culture models for malignant transformation can be used. In 1999 Hahn and Coworkers introduced such a model system and established herewith a basis for research on human tumorigenesis. Primary human fibroblasts are sequentially transduced with defined genetic elements (hTERT, SV40 ER, and H-RasV12), resulting in four defined cell lines, whereby the last has a fully transformed phenotype. In order to get a deeper insight into the molecular biology of human tumorigenesis, we compared the proteomes of these four cell lines following a multimethod concept. At the beginning we assumed SILAC and sample fractionation with COFRADIC is the method of choice to analyze the cell culture model for malignant transformation. Here, the compared samples are combined before sample preparation, thus avoiding differences in sample preparation, and using COFRADIC notably reduces sample complexity. Because 2D-PAGE is a standard method for the separation and visualization of closely related proteomes, we decided to analyze and compare the proteomes of these four cell lines in a first approach by differential 2D-PAGE. Surprisingly, we discovered much more unique results with iTRAQ and sample fractionation with SCX than with the combination of 2D-PAGE and SILAC-COFRADIC. Moreover, iTRAQ outperforms the other strategies not only in number of yielded results but also in analysis time. Here, we present the iTRAQ quantification results and compare them with the results of 2D-PAGE and SILAC-COFRADIC. We found changes in the protein level at each transition. Thereby, SV40 has the strongest impact on the proteome. In detail we identified 201 regulated proteins. Beside others, these proteins are involved in cytoskeleton, RNA processing, and cell cycle, such as CDC2, hnRNPs, snRNPs, collagens, and MCM proteins. For example, MCM proteins are up-regulated and collagens are down-regulated due to SV40 ER expression. Furthermore we made the observation that proteins containing the same domain have analogous regulation profiles during malignant transformation. For instance, several proteins containing a CH or LIM domain are down-regulated. Moreover, by this study and the defined cell culture model, changes could be clearly matched to specific steps during tumorigenesis.

Mineral selective magnetizing flash roasting in oxidizing atmosphere was tested successfully for the improved low intensity magnetic separation of siderite from ankerite and other gangue minerals within ore. The magnetization process was simulated with particle bed tests permeated with artificial combustion gas. The phase transition and the gas composition were measured simultaneously during temperature and time-dependent neutron diffraction experiments. The physical background of the rapid phase transition and the nature of the SO 2 absorption are discussed. Apart from the scientific background, the effects on the process technique are outlined.

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