Enrichment Of Phosphoproteins By Using Guanidine Functionalized Magnetic Materials

Protein phosphorylation is one of the most important post-translational modifications in life activities, which has a closely relationship to the precise regulation of life activities including enzyme activity regulation, neural signals regulation, the proliferation and division of cell, and occur of cancer and so on. A depth recognition of protein phosphorylation such as phosphorylation types, sequences of phosphorylated peptide and phosphorylation sites is very important to understand life activities. The detection and analysis of phosphorylation information is commonly achieved by biological mass spectrometry technique. However, this technology is always hindered by the fact that the ultra low content of phosphorylation protein expressed in living organisms. It has become a necessary opteration to enrich phosphoproteins from complex biological samples prior to mass spectrometry analysis. Thus, preparation of materials capable of selective enrichment of phosphoproteins is significant important to phosphoproteome research.In this study, guanidine functionalized silane was first synthesized, and guanidine functionalized superparamagnetic silica microspheres were synthesized by using the resulting functional monomer. The resulting materials were employed to selectively capture of phosphorylation proteins from complex biological samples, in which FG-1-2 microspheres diplayed the best adsorption ability. FG-1-2 microspheres were characterized by JH NMR, FT-IR, SEM, TEM, TGA, XPS, EDAX, VSM, respectively. The results indicated that FG-1-2 microspheres had a mean size of 280 nm in diameter, the thickness of guanidine-containing silane on microsphere is about 70 nm. Using ?-Casein and ovalbumin as model phosphoproteins, the recognition properties of the obtained materials were evaluated. The adsorption results showed that FG-1-2 microspheres not only has a high adsorption capacity of phosphorylation proteins (63.4 and 74 mg/g for ?-Casein and ovalbumin, respectively) and reach balance in 1 h, but also displayed high selective recognition and enrichment of phosphorylation proteins from complex proteins mixture.The FG-1-2 materials were also used to selectively recognize and enrich of phosphorylation proteins from the actual biological samples such as diluted skim milk and egg white. The results indicated that FG-1-2 displayed high selectivity, which is expected to be further applied to selectively enrich of phosphorylation proteins from other complex actual biological samples.