RESUMO
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.e., as are usually obtained from individual DIGE spots), and that is also applicable to the analysis of complex protein extracts. Additionally, multiple techniques will be described for identifying sites of protein posttranslational modification, with emphasis on phosphorylation and Arg methylation.