Events & Conferences
Book Lists Home
Add To My Book List
Solution isoelectric focusing and its application in comparative proteomic studies of nuclear proteins
ProQuest / UMI
Monday, March 20, 2006
Number of Pages:
In proteomic research, experimental and computational approaches are combined to provide global analysis of the entire proteomes of cells and tissues. The identification and quantification of multiple proteins, which constitute a specific biological system, are important for understanding complex problems in biology. The coupling of highly efficient separations and mass spectrometry instrumentation is evolving rapidly and is being widely applied to problems ranging from biological function to drug development. Development of rapid and high-resolution separation technology is an important field in proteomics. In this study, a solution isoelectric focusing apparatus was modified and built into a two-dimensional separation method for peptides. Newly commercialized isoelectric membranes, which carry immobilized ampholytes, were integrated to establish the pH boundaries in this apparatus. High-performance liquid chromatography was employed as the second dimension, integrated with mass spectrometry. An insoluble nuclear protein fraction was used for optimization and evaluation of this method. The insoluble nuclear proteins were recovered from the nuclei of human MCF-7 human cancer cells and cleaved enzymatically. The resulting peptides were analyzed by the two-dimensional separation method, which coupled solution isoelectric focusing with reversed-phase liquid chromatography interfaced with mass spectrometry. A total of 281 peptides corresponding to 167 proteins were identified by this experiment. The high sample capacity and concentration effect of isoelectric focusing make it possible to detect relatively low abundance proteins in a complex mixture. This two-dimensional separation method dramatically improves peptide detection and identification compared with a single dimension LC-MS analysis. This method has been demonstrated to provide efficient and reproducible separation of both protein and peptides. The two-dimensional separation method was combined with proteolytic isotopic labeling for comparative analysis of protein expression in different cells. Abundances of nuclear proteins from three different drug resistant MCF-7 cancer cell lines were compared to those from the drug susceptible parent cell line using this combined strategy. The abundances of 19 proteins were found to be significantly changed. Their functions are considered in relation to potential mechanisms of in drug resistance.
More Proteomics Books
Methods in Modern Biophysics
Proteomics in Practice: A Laboratory Manual of Proteome Analysis
A comparative proteomic analysis of mitochondrial proteins from drug susceptible and drug resistant human MCF-7 breast cancer cells
Derivatization strategies as an aid to assist separation and mass spectral properties of peptides in proteomics -- Dissertation
Proteomics and Nanocrystallography
Angewandte Bioinformatik: Eine Einführung
Capillary isoelectric focusing-based multidimensional peptide/protein separations for proteomics analysis
The Ten Most Wanted Solutions in Protein Bioinformatics (Chapman & Hall/Crc Mathematical Biology and Medicine)
Proteomics of Spermatogenesis
Microbial Functional Genomics
More Proteomics Books ...
Biohealthmatics.com. All Rights Reserved.
Terms & Conditions
Can't find what you are looking for? View our