Extraction and Solubilization of Proteins for Proteomic Studies

Abstract

For any proteomic study involving various control and experimental specimens, several factors need to be in place. A criticalone is the extraction and solubilization of all components, regardless of whether a chromatographic (1,2) or two-dimensional (2-D) gel electrophoretic fractionation (3–6) is performed prior to analysis of proteins of interest by mass spectrometry of protein digests. All proteins must not onlybe extracted, but they must also be completely soluble, free from interacting partners (such as protein-RNA/DNA and protein-proteininteractions, metabolites, and so on), and, in the case of 2-D gel electrophoresis, they must remain soluble as they approachtheir isoelectric points. The solubilization process should extract all classes of proteins reproducibly, such that statisticallysignificant quantitative data can be obtained and correlated with experimental perturbations and the resulting biologicalresponses.