Identification of proteins based on MS/MS spectra and location of posttranslational modifications.

Comparative protein profiling is a key approach to understanding the human and other proteomes. Systems-level profiling technologies, such as differential fluorescence two-dimensional gel electrophoresis (DIGE), often require the identification of the proteins that are contained within 50 or more spots per gel. A major focus of this chapter therefore is devoted to a general approach for high throughput protein identification that is based on liquid chromatography (LC)/tandem mass spectrometry (MS/MS) analysis of tryptic digests of individual proteins or mixtures of only a few proteins (i.

Proteomics and the analysis of proteomic data: an overview of current protein-profiling technologies.

In recent years, several proteomic methodologies have been developed that now make it possible to identify, characterize, and comparatively quantify the relative level of expression of hundreds of proteins that are coexpressed in a given cell type or tissue, or that are found in biological fluids such as serum. These advances have resulted from the integration of diverse scientific disciplines including molecular and cellular biology, protein/peptide chemistry, bioinformatics, analytical and bioanalytical chemistry, and the use of instrumental and software tools such as multidimensional electrophoretic and chromatographic separations and mass spectrometry. In this unit, some of the common protein-profiling technologies are reviewed, along with the accompanying data-analysis tools.

Ovarian cancer classification based on mass spectrometry analysis of sera.

In our previous study [1], we have compared the performance of a number of widely used discrimination methods for classifying ovarian cancer using Matrix Assisted Laser Desorption Ionization (MALDI) mass spectrometry data on serum samples obtained from Reflectron mode. Our results demonstrate good performance with a random forest classifier. In this follow-up study, to improve the molecular classification power of the MALDI platform for ovarian cancer disease, we expanded the mass range of the MS data by adding data acquired in Linear mode and evaluated the resultant decrease in classification error.

Different endoplasmic reticulum trafficking and processing pathways for calsequestrin (CSQ) and epitope-tagged CSQ.

Cardiac calsequestrin (CSQ) is a protein that traffics to and concentrates inside sarcoplasmic reticulum (SR) terminal cisternae, a protein secretory compartment of uncertain origin. To investigate trafficking of CSQ within standard ER compartments, we expressed CSQ in nonmuscle cell lines and examined its localization by immunofluorescence and its molecular structure from the mass spectrum of total cellular CSQ. In all cells examined, CSQ was a highly phosphorylated protein with a glycan structure predictive of ER-retained proteins: Man9,8GlcNAc2 lacking terminal GlcNAc.