Junying Wei and Yangjun Zhang contributed equally to this work.
Highly efficient enrichment of phosphopeptides by magnetic nanoparticles coated with zirconium phosphonate for phosphoproteome analysis†
Article first published online: 10 MAR 2008
Copyright © 2008 John Wiley & Sons, Ltd.
Rapid Communications in Mass Spectrometry
Volume 22, Issue 7, pages 1069–1080, 15 April 2008
How to Cite
Wei, J., Zhang, Y., Wang, J., Tan, F., Liu, J., Cai, Y. and Qian, X. (2008), Highly efficient enrichment of phosphopeptides by magnetic nanoparticles coated with zirconium phosphonate for phosphoproteome analysis. Rapid Commun. Mass Spectrom., 22: 1069–1080. doi: 10.1002/rcm.3485
- Issue published online: 26 MAR 2008
- Article first published online: 10 MAR 2008
- Manuscript Accepted: 3 FEB 2008
- Manuscript Revised: 30 JAN 2008
- Manuscript Received: 31 OCT 2007
- National Natural Science Foundation of China. Grant Numbers: 20505018, 20505019, 20635010, 20735005, 30621063
- National Key Program for Basic Research of China. Grant Numbers: 2006CB910801, 2006CB910803, 2007CB714104, 2006AA02A308
The location of phosphorylation plays a vital role for the elucidation of biological processes. The challenge of low stoichiometry of phosphoproteins and signal suppression of phosphopeptides by nonphosphopeptides in mass spectrometry (MS) analysis makes the selective enrichment of phosphopeptides prior to MS analysis necessary. Besides the immobilized metal affinity chromatography (IMAC) method, some affinity methods based on nanoparticles displayed a higher enrichment efficiency for phosphopeptides such as Fe3O4/TiO2 and Fe3O4/ZrO2 nanoparticles. To further improve the selectivity and compatibility of the affinity methods, a novel strategy based on magnetic nanoparticles coated with zirconium phosphonate for the enrichment of phosphopeptides has been developed in this study. Under optimized experimental conditions, 1 × 10−9 M phosphopeptides in 50 µL tryptic digest of β-casein could be enriched and identified successfully. Reliable results were also obtained for 1 × 10−8 M phosphopeptides in 50 µL tryptic digest of β-casein in the presence of nonphosphopeptides from a tryptic digest of bovine serum albumin (BSA) over 20 times in concentration. The performance of nanoparticles for use in a real sample was further demonstrated by employing the strong cation-exchange chromatography (SCX) fraction of a tryptic digest of a protein extract from Chang liver cells as a model sample. Experimental results show that the nanoparticles can be easily and effectively used for enrichment of phosphopeptides in low concentration. Most importantly, our approach is more compatible with commonly used SCX strategies than Fe3+-IMAC. The proposed method thus has great potential for future studies of large-scale phosphoproteomes. Copyright © 2008 John Wiley & Sons, Ltd.