Functionalized carbon encapsulated iron nanoparticles as affinity probes for enrichment of phosphoproteins /phosphopeptides

• Main Authors: Huang-juo Chen, 陳煌焯
• Other Authors: Chih-Che Wu
• Format: Others
• Language: zh-TW
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/93544464646933115029

碩士 === 國立暨南國際大學 === 應用化學系 === 97 === Abstract Carbon encapsulated magnetic nanoparticles have wide applications such as, serving as contrast reagents for magnetic resonance imaging (MRI), magnetic data storage medium, and magnetic hyperthermia. However, compared to the intense studies in the synthetic strategies for fabricating such core/shell materials, little effort has been devoted to the surface functionalziation strategies. In the report, we produced polyarginine-immobilized and titanium dioxide-immobilized carbon encapsulated magnetic iron nanoparticles (Fe@CNPs) as high affinity probes to extract and enrich phosphopeptides and phosphoprotein from a number of enzymatic digested products including α- and β-casein and milk sample. The poly(arcylic acid)-coated Fe@CNPs can be easily grafted with affinity ligands such as polyarginine (PA) or titanium dioxide via carbodiimide chemistry or sol-gel chemistry, respectively. For the capture of phosphopeptides, only a 10-s incubation time with affinity probes is sufficient to prove faithful information. The affinity probes with trapped phosphopeptides are then isolated by magnetic separation, cleaned with washing solvents to remove nonspecifically bound peptides. Finally, the trapped phosphopeptides were eluted with 2,5-dihydroxybenzoic acid (2,5-DHB) containing 5% phosphoric acid and deposited onto the MALDI target plate for analysis by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). After enrichment, the ion suppression caused by abundant nonphosphopeptides is eliminated, and the detection sensitivity of phosphopeptides by MALDI MS analysis is in the low femtomol range Both polyarginine and TiO2-coated Fe@CNPs exhibits good specificity and sensitivity toward phosphopeptides. These two types of affinity probes process complementary properties for phosphopeptide mapping by MALID MS and can be combined to provide rich information for phosphoproteomic analysis.