MASS SPECTROSCOPY/PROTEOMICS

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DESCRIPTION (provided by applicant): The Mass Spectrometry/Proteomics Shared Facility is organized to provide a variety of analytical and technical services using mass spectrometry to UAB Comprehensive Cancer Center investigators. The Shared Facility currently has five mass spectrometers - two MALD1-TOF instruments, two triple quadrupole instruments, and one hybrid quadrupole orthogonal TOF instrument. Three new instruments (a new triple quadrupole instrument and two fourier transform-ion cyclotron resonance instruments - supported by NCRR grants) will be installed during the summer of 2004. Besides qualitative and quantitative analysis of small molecules (therapeutic and xenobiotics) and peptides, the mass spectrometry group provides a variety of analyses in the area of protein chemistry and protein expression. These include H-D exchange to probe protein structure in solution as well as protein-protein interactions. Procedures have also recently been added to allow investigators to pursue proteomics applications. The proteomics section of the Shared Facility will provide investigators with consultant advice as how to prepare samples for analysis, and in most cases to carry out the analyses. This includes recovery of the proteome from the cell or tissue samples, resolving proteins using 1D-SDS-PAGE or 2D-IEF/SDS-PAGE, staining and imaging the gels, and carrying out sophisticated image analysis to detect changes that have occurred in the observable proteome. These are verified by statistical analysis by the Biostatistics/Bioinformatics Shared Facility. Protein spots are removed robotically and following robotic processing to generate a protease-induced peptide profile analysis by MALDI-TOF mass spectrometry. The peptide mass fingerprint is subjected to statistical analysis with the MASCOT search engine to putatively identify the protein. Verification of protein identity is performed by nano-LC tandem mass spectrometry of the peptides. This method also allows for the detection of post-translational modifications.