Preparation Of TiO₂-MWNTs By Hydrothermal Reaction Process To Determine Phosphopeptides

Protein phosphorylation is one of the most important post-translational modifications in cells, which regulating of a wide variety of cellular events including cell division, growth, migration, differentiation and signal transduction. Due to the central regulation role of phosphorylation, so the investigation and study of phosphoproteins and the conforming of phosphorylation sites are very important in proteome. Currently, mass spectrometry (MS) based techniques are the firstly choice in the structure analysis of phosphopeptides. However, because of the substoichiometric and difficult ionization of phosphopeptides, the study of protein phosphorylation by using MS is still a big challenge. So the enrichment of phosphoproteins and phosphopeptides before MS analysis is one of the important steps to improve the MS response of phosphopeptide signals.In this study, titanium dioxide-multi walled carbon nanotubes (TiO2-MWNTs) nanocomposite synthesized by hydrothermal reaction process. The prepared TiO2-MWNTs nanocomposite was characterized by SEM TEM, FTIR and XRD, and the results demonstrated that the anatase phase titanium dioxide (TiO2) nanoparticles had been successfully linked with MWCNTs. TiO2nanoparticles have a mean diameter10nm and the crystal formation is anatase mostly. TiO2-MWNTs nanocomposite was used to selectively enrich phosphopeptides from β-casein digest and to estimate the effect enrichment. Optimization of the enrichment conditions, the sample liquid organic solvent concentration of60%acetonitrile, the concentration of trifluoroacetic acid is0.1%, solution ionic strength was controlled by sodium chloride, nonphosphopeptides are inhibited by lactic acid, washing times of twice and elution as0.4mol/L. The results show that this method has high selectivity phosphorylated peptides. Enrichment of phosphopeptides from the tryptic digest of β-casein/BSA in the molar ratios of1:50and1:100, respectively, it is clear that TiO2-MWNTs can specifically and selectively bind phosphopeptides from the complex sample solution. This nanocomposite material was further applied to enrich the phosphopeptides in a protein digest obtained from nonfat milk successfully.