Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Department of Chemistry, Wuhan University, Wuhan 430072, China.
J Chromatogr A. 2013 Nov 8;1315:61-9. doi: 10.1016/j.chroma.2013.09.057. Epub 2013 Sep 19.
As one of the most important post-translational modifications, reversible phosphorylation of protein plays crucial roles in a large number of biological processes. Moreover, endogenous phosphopeptides are also associated with certain human diseases. An efficient enrichment and separation method is the premise for successful identification and quantification of phosphopeptides. In this work, titanium grafted magnetic mesoporous silica (Fe3O4@Ti-mSiO2) was developed and applied for the enrichment of endogenous phosphopeptides. Fe3O4@Ti-mSiO2 particles were prepared by grafting titanocene dichloride on the inner walls of magnetic mesoporous silica and then being calcined to remove cyclopentadienyl ligand. The physicochemical properties of the prepared materials were characterized by energy dispersive X-ray spectroscopy (EDX), nitrogen adsorption-desorption analysis, X-ray diffraction (XRD), transmission electron microscopy (TEM), scanning electron microscopy (SEM) and vibrating sample magnetometer (VSM). For selective enrichment of phosphopeptides, the prepared Fe3O4@Ti-mSiO2 particles were applied for tryptic digests of β-casein, mixtures of β-casein and bovine serum albumin (BSA), and low-fat milk. Finally, Fe3O4@Ti-mSiO2 was successfully applied for the enrichment of endogenous phosphopeptides from human serum.
作为最重要的翻译后修饰之一，蛋白质的可逆磷酸化在大量的生物过程中起着至关重要的作用。此外，内源性磷酸肽也与某些人类疾病有关。高效的富集和分离方法是成功鉴定和定量磷酸肽的前提。在这项工作中，开发了钛接枝磁性介孔硅（Fe3O4@Ti-mSiO2）并将其用于内源性磷酸肽的富集。通过将二氯二茂钛接枝到磁性介孔硅的内壁上，然后煅烧以去除环戊二烯基配体，制备了 Fe3O4@Ti-mSiO2 颗粒。通过能量色散 X 射线光谱（EDX）、氮气吸附-解吸分析、X 射线衍射（XRD）、透射电子显微镜（TEM）、扫描电子显微镜（SEM）和振动样品磁强计（VSM）对制备材料的物理化学性质进行了表征。为了选择性地富集磷酸肽，将制备的 Fe3O4@Ti-mSiO2 颗粒应用于β-酪蛋白的酶解物、β-酪蛋白和牛血清白蛋白（BSA）的混合物以及低脂牛奶。最后，Fe3O4@Ti-mSiO2 成功地应用于从人血清中富集内源性磷酸肽。
