Stable isotope phospho-profiling of fibrinogen and fetuin subunits by element mass spectrometry coupled to capillary liquid chromatography.
We have used one-dimensional polyacrylamide gel electrophoresis, tryptic digestion, and capillary liquid chromatography-mass spectrometry with inductively coupled plasma ionization and phosphorus-31 detection or electrospray ionization for the analysis of protein phosphorylation. We have analyzed human fibrinogen with two well-characterized phosphorylation sites and bovine fetuin with unknown phosphorylation status. Both serine-3 and serine-345 (both in Aalpha) of fibrinogen were clearly recognized. In bovine fetuin, four phosphorylated sites were newly characterized (serine-138, serine-320, serine-323, and serine-324). The novel strategy provides a fast and quantitative overview of the presence of protein phosphorylation sites.
Central Spectroscopy, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
Mathias
M
Wind
Darko
D
Gosenca
Dieter
D
Kübler
Wolf D
WD
Lehmann
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D019075:Electrophoresis, Capillary / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D005340:Fibrinogen / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis; D010759:Phosphorus Isotopes; D010766:Phosphorylation; D021122:Protein Subunits; D012694:Serine / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D000509:alpha-Fetoproteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism
D010446:Peptide Fragments; D010759:Phosphorus Isotopes; D021122:Protein Subunits; D000509:alpha-Fetoproteins; D012694:Serine; D005340:Fibrinogen