Gel-free proteomics

Advantages and Applications of Gel-Free Proteomic Approaches in the Study of Prokaryotes... 

STATE OF AFFAIRS IN THE STUDY OF PROKARYOTES VIA GEL-FREE PROTEOMICS... 

In the last years the application of two-dimensional electrophoresis (2-DE) has often been declared outdated and a new century of gel-free proteomics was announced. Nevertheless, 2-DE is still the method of choice when analyzing complex protein mixtures. With a separation of10 000 proteins, 2-DE gives access to high-resolution proteome analysis. Continuous development has consolidated 2-DE application in proteomics, where the introduction of difference gel electrophoresis (DICE) is the latest improvement. DICE is based on fluorescently tagging all proteins in each sample with one set of matched fluorescent dyes designed to minimally interfere with protein mobility during 2-DE. 

K. Gevaert, J. Van Damme, M. Goethals, G.R. Thomas, B. Hoorelbeke, H. Demol, L. Martens, M. Puype, A. Staes, J. Vandekerckhove, Chromatographic isolation of Met-containing peptides for gel-free proteome analysis Identif cation of more than 800 Escherichia coliproteins. Mol Cell Proteomics 1 (2002) 896. 

Gevaert K, Van Damme J, Goethals M, et al. (2002). Chromatographic isolation of methionine-containing peptides for gel-free proteome analysis Identification of more than 800 Escherichia coli proteins. Mol. Cell Proteomics. l(ll) 896-903. 

The membrane-bound cytochrome P450 enzymes represent key players in secondary metabolite pathways as they catalyze vital, often rate-limiting steps. However, basic, hydro-phobic and membrane-spanning proteins remain an intrinsic limitation for 2D-PAGE [118]. Adoption of gel free proteomic approaches such as iTRAQ (Table 6) may help to overcome this bottleneck. 

In gel-free proteomic approaches, also named shotgun proteomics, the entire proteome of a biological system is enzymatically digested and the very complex peptide mixture thus obtained, containing thousands of peptides, is directly analyzed by two-or multi-dimensional LC-MS/MS... 

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