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Phosphoproteome Analysis

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Bioscience Reports

Abstract

Protein phosphorylation is directly or indirectly involved in all important cellular events. The understanding of its regulatory role requires the discovery of the proteins involved in these processes and how, where and when protein phosphorylation takes place. Investigation of the phosphoproteome of a cell is becoming feasible today although it still represents a very difficult task especially if quantitative comparisons have to be made. Several different experimental strategies can be employed to explore phosphoproteomes and this review will cover the most important ones such as incorporation of radiolabeled phosphate into proteins, application of specific antibodies against phosphorylated residues and direct staining of phosphorylated proteins in polyacrylamide gels. Moreover, methods to enrich phosphorylated proteins such as affinity chromatography (IMAC) and immunoprecipitation as well as mass spectrometry for identification of phosphorylated peptides and phosphorylation sites are also described.

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Abbreviations

IMAC:

immobilized metal affinity chromatography

1D:

one dimensional

2D:

two dimensional

ECL:

enhanced chemiluminescence

pY:

phosphotyrosine

SDS-PAGE:

sodium dodecyl sulfate polyacrylamide gel electrophoresis

MP:

multiplex proteomics

DIGE:

difference gel electrophoresis

SILAC:

stable isotope labeling with amino acids in cell culture

CID:

collision-induced dissociation

amu:

atomic mass unit

ESI:

electrospray ionization

λ PPase:

lambda phosphatase

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  1. Sienabiotech S.p.A., Discovery Research, Via Fiorentina 1, 53100, Siena, Italy

    Roberto Raggiaschi, Stefano Gotta & Georg C. Terstappen

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  1. Roberto Raggiaschi
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  2. Stefano Gotta
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  3. Georg C. Terstappen
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Correspondence to Roberto Raggiaschi.

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Raggiaschi, R., Gotta, S. & Terstappen, G.C. Phosphoproteome Analysis. Biosci Rep 25, 33–44 (2005). https://doi.org/10.1007/s10540-005-2846-0

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