Analysis of protein phosphorylation using mass spectrometry.

Protein phosphorylation has been known to be a pivotal modification regulating many cellular activities and functions. Except for several conventional techniques, mass spectrometry-based strategies are increasingly considered as vital tools that can be utilized to characterize phosphorylated peptides or proteins. In this article, we summarized currently available mass spectrometry-based techniques for the analysis of phosphorylation. Due to the low abundance of phosphopeptides, enrichment steps such as specific antibodies, immobilized metal affinity chromatography, and specific tags are crucial for their use in detection. Since the non-specific binding of the enrichment techniques are constantly of major concerns, phosphatase treatment, neutral loss scan, or precursor ion scan enable the recognition of the phosphopeptide signals. In addition, quantitative methods including isotope labeling and mass tags are also discussed. Phosphoproteome analysis seems to provide elucidation of signaling networks and global decipherment of cell activities, which require powerful analytical methods for complete and routine identification of the phosphorylation event. Despite that numerous approaches have been exploited, comprehensive analysis of protein phosphorylation remains a challenging task. With the progressively more improvements of instruments and methodologies, we can foresee the implementation of a comprehensive approach for the analysis of phosphorylation states of proteins.

[1]  Hsin-Yi Wu,et al.  Mining phosphopeptide signals in liquid chromatography-mass spectrometry data for protein phosphorylation analysis. , 2007, Journal of proteome research.

[2]  Ken Aoshima,et al.  Enhancement of the efficiency of phosphoproteomic identification by removing phosphates after phosphopeptide enrichment. , 2007, Journal of proteome research.

[3]  W. Weckwerth,et al.  Enrichment of phosphorylated proteins and peptides from complex mixtures using metal oxide/hydroxide affinity chromatography (MOAC) , 2005, Proteomics.

[4]  Yu-Chie Chen,et al.  Fe3O4/TiO2 core/shell nanoparticles as affinity probes for the analysis of phosphopeptides using TiO2 surface-assisted laser desorption/ionization mass spectrometry. , 2005, Analytical chemistry.

[5]  C. Borchers,et al.  Phosphatase-directed phosphorylation-site determination: a synthesis of methods for the detection and identification of phosphopeptides. , 2005, Journal of proteome research.

[6]  D. Lauffenburger,et al.  Time-resolved Mass Spectrometry of Tyrosine Phosphorylation Sites in the Epidermal Growth Factor Receptor Signaling Network Reveals Dynamic Modules*S , 2005, Molecular & Cellular Proteomics.

[7]  A. Harmon,et al.  Enhancement of phosphoprotein analysis using a fluorescent affinity tag and mass spectrometry. , 2005, Rapid communications in mass spectrometry : RCM.

[8]  Achim Kramer,et al.  Mapping of phosphorylation sites by a multi-protease approach with specific phosphopeptide enrichment and NanoLC-MS/MS analysis. , 2005, Analytical chemistry.

[9]  P. Roepstorff,et al.  Highly Selective Enrichment of Phosphorylated Peptides from Peptide Mixtures Using Titanium Dioxide Microcolumns* , 2005, Molecular & Cellular Proteomics.

[10]  Benjamin A Garcia,et al.  Analysis of protein phosphorylation by mass spectrometry. , 2005, Methods.

[11]  M. Mann,et al.  Quantitative Phosphoproteomics Applied to the Yeast Pheromone Signaling Pathway*S , 2005, Molecular & Cellular Proteomics.

[12]  M. Gustafsson,et al.  Detection of phosphorylated peptides in proteomic analyses using microfluidic compact disk technology. , 2004, Analytical chemistry.

[13]  A. Heck,et al.  Selective isolation at the femtomole level of phosphopeptides from proteolytic digests using 2D-NanoLC-ESI-MS/MS and titanium oxide precolumns. , 2004, Analytical chemistry.

[14]  Angela Bachi,et al.  Analysis of Protein Phosphorylation by Mass Spectrometry , 2004, European journal of mass spectrometry.

[15]  Matthias Mann,et al.  A Mass Spectrometry-based Proteomic Approach for Identification of Serine/Threonine-phosphorylated Proteins by Enrichment with Phospho-specific Antibodies , 2002, Molecular & Cellular Proteomics.

[16]  Hanno Steen,et al.  Analysis of protein phosphorylation using mass spectrometry: deciphering the phosphoproteome. , 2002, Trends in biotechnology.

[17]  J. Shabanowitz,et al.  Phosphoproteome analysis by mass spectrometry and its application to Saccharomyces cerevisiae , 2002, Nature Biotechnology.

[18]  Richard D. Smith,et al.  Phosphoprotein isotope-coded affinity tags: application to the enrichment and identification of low-abundance phosphoproteins. , 2002, Analytical chemistry.

[19]  B. Chait,et al.  Analysis of phosphorylated proteins and peptides by mass spectrometry. , 2001, Current opinion in chemical biology.

[20]  H. Steen,et al.  Quadrupole time-of-flight versus triple-quadrupole mass spectrometry for the determination of phosphopeptides by precursor ion scanning. , 2001, Journal of mass spectrometry : JMS.

[21]  I. Kerr,et al.  Phosphotyrosine profiling to identify novel components of interferon and interleukin 6‐family cytokine signalling , 2001, Proteomics.

[22]  Richard D. Smith,et al.  Phosphoprotein isotope-coded affinity tag approach for isolating and quantitating phosphopeptides in proteome-wide analyses. , 2001, Analytical chemistry.

[23]  R. Aebersold,et al.  A systematic approach to the analysis of protein phosphorylation , 2001, Nature Biotechnology.

[24]  B. Chait,et al.  Enrichment analysis of phosphorylated proteins as a tool for probing the phosphoproteome , 2001, Nature Biotechnology.

[25]  A. Pandey,et al.  Detection of tyrosine phosphorylated peptides by precursor ion scanning quadrupole TOF mass spectrometry in positive ion mode. , 2001, Analytical chemistry.

[26]  A. Stensballe,et al.  Characterization of phosphoproteins from electrophoretic gels by nanoscale Fe(III) affinity chromatography with off‐line mass spectrometry analysis , 2001, Proteomics.

[27]  P. Roepstorff,et al.  Phospho‐proteomics: Evaluation of the use of enzymatic de‐phosphorylation and differential mass spectrometric peptide mass mapping for site specific phosphorylation assignment in proteins separated by gel electrophoresis , 2001, Proteomics.

[28]  W. Lehmann,et al.  Analysis of protein phosphorylation by a combination of elastase digestion and neutral loss tandem mass spectrometry. , 2001, Analytical chemistry.

[29]  H. Lodish,et al.  Identification of a Novel Immunoreceptor Tyrosine-based Activation Motif-containing Molecule, STAM2, by Mass Spectrometry and Its Involvement in Growth Factor and Cytokine Receptor Signaling Pathways* , 2000, The Journal of Biological Chemistry.

[30]  P. Cohen,et al.  The regulation of protein function by multisite phosphorylation--a 25 year update. , 2000, Trends in biochemical sciences.

[31]  O. Fiehn,et al.  Comparative quantification and identification of phosphoproteins using stable isotope labeling and liquid chromatography / mass spectrometry , 2022 .

[32]  K. Tomer,et al.  Detection and sequencing of phosphopeptides , 2000, Journal of the American Society for Mass Spectrometry.

[33]  T. Hunter,et al.  Signaling—2000 and Beyond , 2000, Cell.

[34]  M. Mann,et al.  Analysis of receptor signaling pathways by mass spectrometry: identification of vav-2 as a substrate of the epidermal and platelet-derived growth factor receptors. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[35]  M. Quadroni,et al.  Phosphopeptide analysis. , 2000, EXS.

[36]  M. Posewitz,et al.  Immobilized gallium(III) affinity chromatography of phosphopeptides. , 1999, Analytical chemistry.

[37]  F. Cross,et al.  Accurate quantitation of protein expression and site-specific phosphorylation. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[38]  A. J. McQuillan,et al.  Phosphate Adsorption onto TiO2 from Aqueous Solutions: An in Situ Internal Reflection Infrared Spectroscopic Study , 1999 .

[39]  N. Packer,et al.  Protein phosphorylation: technologies for the identification of phosphoamino acids. , 1998, Journal of chromatography. A.

[40]  A. Hinnebusch,et al.  Identification of phosphorylation sites in proteins separated by polyacrylamide gel electrophoresis. , 1998, Analytical chemistry.

[41]  S. Carr,et al.  Selective detection and sequencing of phosphopeptides at the femtomole level by mass spectrometry. , 1996, Analytical biochemistry.

[42]  G. Neubauer,et al.  Parent ion scans of unseparated peptide mixtures. , 1996, Analytical chemistry.

[43]  A. P. Hunter,et al.  Chromatographic and mass spectrometric methods for the identification of phosphorylation sites in phosphoproteins. , 1994, Rapid communications in mass spectrometry : RCM.

[44]  J Allison,et al.  An approach to locate phosphorylation sites in a phosphoprotein: mass mapping by combining specific enzymatic degradation with matrix-assisted laser desorption/ionization mass spectrometry. , 1994, Analytical biochemistry.

[45]  M. F. Bean,et al.  Selective detection of phosphopeptides in complex mixtures by electrospray liquid chromatography/mass spectrometry , 1993, Journal of the American Society for Mass Spectrometry.

[46]  P. Cohen,et al.  On target with a new mechanism for the regulation of protein phosphorylation. , 1993, Trends in biochemical sciences.

[47]  L. Heilmeyer,et al.  Sequence analysis of phosphoserine‐containing peptides , 1986, FEBS letters.

[48]  J. Porath,et al.  Isolation of phosphoproteins by immobilized metal (Fe3+) affinity chromatography. , 1986, Analytical biochemistry.

[49]  J. A. Nimmo,et al.  The identification of phosphoseryl residues during the determination amino acid sequence in phosphoproteins. , 1982, Analytical biochemistry.