Relative quantification of erythropoietin receptor-dependent phosphoproteins using in-gel 18O-labeling and tandem mass spectrometry.
On examining different proteomics approaches for the investigation of structure-function relationships of erythropoietin (EPO) receptor signaling, it was found that two-dimensional gel electrophoresis/mass spectrometry procedures are clearly limited in their ability to detect low-expressed signaling proteins. Instead it was found that a strategy involving anti-phosphotyrosine immunoprecipitation, one-dimensional gel electrophoresis (1DE), and capillary liquid chromatography/tandem mass spectrometry (LC/MS/MS) provides the sensitivity required for identification of signaling proteins. In the present work the immunoprecipitation/1DE/LC/MS approach was combined with an in-gel 18O-labeling technique to analyze EPO receptor-dependent proteins. Identification and relative quantification of more than 180 EPO receptor-dependent proteins were achieved directly based on the in-gel 18O-labeling approach.
Körbel
Sandra
S
Institut für Medizinische Biochemie und Molekularbiologie, Universität Rostock, 18057 Rostock, Germany.
Schümann
Michael
M
Bittorf
Thomas
T
Krause
Eberhard
E
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D010103:Oxygen Isotopes; D010750:Phosphoproteins / Q000032:analysis* / Q000737:chemistry*; D017467:Receptors, Erythropoietin / Q000378:metabolism*; D012015:Reference Standards; D012680:Sensitivity and Specificity
D010103:Oxygen Isotopes; D010750:Phosphoproteins; D017467:Receptors, Erythropoietin