Mapping the phosphorylation sites of proteins using on-line immobilized metal affinity chromatography/capillary electrophoresis/electrospray ionization multiple stage tandem mass spectrometry
✍ Scribed by Ping Cao; John T. Stults
- Publisher
- John Wiley and Sons
- Year
- 2000
- Tongue
- English
- Weight
- 127 KB
- Volume
- 14
- Category
- Article
- ISSN
- 0951-4198
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✦ Synopsis
On
-line immobilized metal affinity chromatography/capillary electrophoresis/electrospray ionization-mass spectrometry (IMAC/CE/ESI-MS) offers selective preconcentration of phosphorylated peptides with identification of the phosphorylated amino acid(s). The preconcentration provides low concentration limits of detection and capillary electrophoresis separates the peptides. Recently, we reported a fast, simple, and sensitive on-line IMAC/CE/ESI-MS/MS method for the determination of phosphopeptides at low-pmole levels. That work is expanded here by use of multiple stage tandem mass spectrometry (MS n , n = 2,3) to isolate and fragment target ions to provide more reliable assignments of phosphorylated residues. The application of IMAC/CE/ESI-MS n is demonstrated by the analysis of tryptic digests of a-and b-casein and in-gel tryptic digests of b-casein.