Chapter 9: Proteomic Methods for Phosphorylation Site Mapping—References

Abu-lawi K.I. and Sultzer B.M. 1995. Induction of serine and threonine protein phosphorylation by endotoxin-associated protein in murine resident peritoneal macrophages. Infect. Immun. 63: 498–502.

Adams J.A. 2001. Kinetic and catalytic mechanisms of protein kinases. Chem. Rev. 101: 2271–2290.

Aebersold R. and Goodlett D. 2001. Mass spectrometry in proteomics. Chem. Rev. 101: 269–295.

Aebersold R., Watts J.D., Morrison H.D., and Bures E. 1991. Determination of the site of tyrosine phosphorylation at the low picomole level by automated solid-phase sequence analysis. Anal. Biochem. 199: 51–60.

Aebersold R., Figeys D., Gygi S., Corthals G., Haynes P., Rist B., Zhang Y., and Goodlett D.R. 1998. Towards an integrated analytical technology for the generation of multidimensional protein expression maps. J. Prot. Chem. 17: 533–535.

Affolter M., Watts J.D., Krebs D.L., and Aebersold R. 1994. Evaluation of two-dimensional phosphopeptide maps by electrospray ionization mass spectrometry of recovered peptides. Anal. Biochem. 223: 74–81.

Ahn N.G. and Resing K.A. 2001. Toward the phosphoproteome. Nat. Biotechnol. 19: 317–318.

Amankwa L.N., Harder K., Jirik F., and Aebersold R. 1995. High-sensitivity determination of tyrosine-phosphorylated peptides by on-line enzyme reactor and electrospray ionization mass spectrometry. Protein Sci. 4: 113–125.

Andersson L. and Porath J. 1986. Isolation of phosphoproteins by immobilized metal (Fe³⁺) affinity chromatography. Anal. Biochem. 154: 250–254.

Annan R.S. and Carr S.A. 1996. Phosphopeptide analysis by matrix-assisted laser desorption time-of-flight mass spectrometry. Anal. Chem. 68: 3413–3421.

^_______. 1997. The essential role of mass spectrometry in characterizing protein structure: Mapping posttranslational modifications. J. Protein Chem. 16: 391–402.

Annan R.S., Huddleston M.J., Verma R., Deshaies R.J., and Carr S.A. 2001. A multidimensional electrospray MS-based approach to phosphopeptide mapping. Anal. Chem. 73: 393–404.

Arad-Dann H., Beller U., Haimovitch R., Gavrieli Y., and Ben-Sasson S.A. 1993. Immunohistochemistry of phosphotyrosine residues: Identification of distinct intracellular patterns in epithelial and steroidogenic tissues. J. Histochem. Cytochem. 41: 513–519.

Asara J.M. and Allison J. 1999. Enhanced detection of phosphopeptides in matrix-assisted laser desorption/ionization mass spectrometry using ammonium salts. J. Am. Soc. Mass Spectrom. 10: 35–44.

Baldwin M.A., Medzihradszky K.F., Lock C.M., Fisher B., Settineri T.A., and Burlingame A.L. 2001. Matrix-assisted laser desorption/ionization coupled with quadrupole/orthogonal acceleration time-of-flight mass spectrometry for protein discovery, identification, and structural analysis. Anal. Chem. 73: 1707– 1720.

Bateman R.H., Carruthers R., Hoyes J.B., Jones C., Langridge J.I., Millar A., and Vissers J.P. 2002. A novel precursor ion discovery method on a hybrid quadrupole orthogonal acceleration time-of-flight (Q-TOF) mass spectrometer for studying protein phosphorylation. J. Am. Soc. Mass Spectrom. 13: 792–803.

Becker S., Corthals G.L., Aebersold R., Groner B., and Muller C.W. 1998. Expression of a tyrosine phosphorylated, DNA binding Stat3beta dimer in bacteria. FEBS Lett. 441: 141–147.

Bennett K.L., Stensballe A., Podtelejnikov A., Moniatte M., and Jensen O.N. 2000. Phosphopeptide analysis using a MALDI-QqTOF mass spectrometer. In Proceedings of the 48th ASMS Conference on Mass Spectrometry and Allied Topics, pp. 627–628. American Society for Mass Spectrometry, Long Beach, California.

^_______. 2002. Phosphopeptide detection and sequencing by matrix-assisted laser desorption/ionization quadrupole time-of-flight tandem mass spectrometry. J. Mass Spectrom. 37: 179–190.

Benore-Parsons M., Seidah N.G., and Wennogle L.P. 1989. Substrate phosphorylation can inhibit proteolysis by trypsin-like enzymes. Arch. Biochem. Biophys. 272: 274–280.

Biemann K. 1988. Contributions of mass spectrometry to peptide and protein structure. Biomed. Environ. Mass Spectrom. 16: 99–111.

^_______. 1992. Mass spectrometry of peptides and proteins. Annu. Rev. Biochem. 61: 977–1010.

Blume-Jensen P. and Hunter T. 2001a. Two-dimensional phosphoamino acid analysis. Methods Mol. Biol. 124: 49–65.

^_______. 2001b. Oncogenic kinase signalling. Nature 411: 355–365.

Bond J.A., Webley K., Wyllie F.S., Jones C.J., Craig A., Hupp T., and Wynford-Thomas D. 1999. p53-Dependent growth arrest and altered p53-immunoreactivity following metabolic labeling with ³²P ortho-phosphate in human fibroblasts. Oncogene 18: 3788–3792.

Boyle W.J., Geer Van der P., and Hunter T. 1991. Phosphopeptide mapping and phosphoamino acid analysis by two-dimensional separation on thin-layer cellulose plates. Methods Enzymol. 201: 110–149.

Cao P. and Stults J.T. 1999. Phosphopeptide analysis by on-line immobilized metal-ion affinity chromatography-capillary electrophoresis electrospray ionization mass spectromtry. J. Chromatogr. A 853: 225–235.

^_______. 2000. Mapping the phosphorylation sites of proteins using on-line immobilized metal affinity chromatography/capillary electrophoresis/electrospray ionization multiple stage tandem mass spectrometry. Rapid Commun. Mass Spectrom. 14: 1600–1606.

Carr S.A., Huddleston M.J., and Annan R.S. 1996. Selective detection and sequencing of phosphopeptides at the femtomole level by mass spectrometry. Anal. Biochem. 239: 180–192.

Chen J., Qi Y., Zhao R., Zhou G.W., and Zhao Z.J. 2001. Assay of protein tyrosine phosphatases by using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Anal. Biochem. 292: 51– 58.

Chen S.L., Huddleston M.J., Shou W., Deshaies R.J., Annon R., and Carr S.A. 2002. Mass spectrometry-based methods for phosphorylation site mapping of hyperphosphorylated proteins applied to Net1, a regulator of exit from mitosis in yeast. Mol. Cell. Proteomics 1: 186–196.

Cleverley K.E., Betts J.C., Blackstock W.P., Gallo J.-M., and Anderton B. H. 1998. Identification of novel in vitro pka phosphorylation sites on the low and middle molecular mass neurofilament subunits by mass spectrometry. Biochemistry 37: 3917–3930.

Cohen P. 2002. The origins of protein phosphorylation. Nat. Cell Biol. 4: E127–130.

Conrads T.P., Issaq H.J., and Veenstra T.D. 2002. New tools for quantitative phosphoproteome analysis. Biochem. Biophys. Res. Commun. 290: 885–890.

Corthals G.L., Gygi S.P., Aebersold R., and Patterson S.D. 1999. Identification of proteins by mass spectrometry. In Proteome research: 2D gel electrophoresis and detection methods (ed. T. Rabilloud), pp. 197–231. Springer, New York.

Covey T.R., Huang E.C., and Henion J.D. 1991a. Structural characterization of protein tryptic peptides via liquid chromatography/mass spectrometry and collision-induced dissociation of their doubly charged molecular ions. Anal. Chem. 63: 1193–2000.

Covey T., Shushan B., Bonner R., Schröder W., and Hucho F. 1991b. LC/MS and LC/MS/MS screening for the sites of post-translational modifications in proteins. In Methods in protein sequence analysis (ed. H. Jörnvall et al.), pp. 249–256. Birkhäuser Verlag, Basel, Switzerland.

Craig A.G. 2001. Identification of the sites of phosphorylation in proteins using high performance liquid chromatography and mass spectrometry. Methods Mol. Biol. 124: 87–105.

Edman P. 1949. A method for the determination of the amino acid sequence in peptides. Arch. Biochem. 22: 475–476.

Fenn J.B., Mann M., Meng C.K., Wong S.F., and Whitehouse C.M. 1989. Electrospray ionization for mass spectrometry of large biomolecules. Science 246: 64–71.

Figeys D., Corthals G.L., Gallis B., Goodlett D.R., Ducret A., Corson M.A., and Aebersold R. 1999. Data-dependent modulation of solid-phase extraction capillary electrophoresis for the analysis of complex peptide and phosphopeptide mixtures by tandem mass spectrometry: Application to endothelial nitric oxide synthase. Anal. Chem. 71: 2279–2287.

Flora J.W. and Muddiman D.C. 2001. Selective, sensitive, and rapid phosphopeptide identification in enzymatic digests using ESI-FTICR-MS with infrared multiphoton dissociation. Anal. Chem. 73: 3305–3311.

Francis S.H. and Corbin J.D. 1993. Structure and function of cyclic nucleotide-dependent protein kinases. Ann. Rev. Physiol. 56: 237–272.

Gallis B., Corthals G.L., Goodlett D.R., Ueba H., Kim F., Presnell S.R., Figeys D., Harrison D.G., Berk B.C., Aebersold R., and Corson M. 1999. Identification of flow-dependent endothelial nitric-oxide synthase phosphorylation sites by mass spectrometry and regulation of phosphorylation and nitric oxide production by the phosphatidylinositol 3-kinase inhibitor LY294002. J. Biol. Chem. 274: 30101–30108.

Garnier C., Lafitte D., Jorgensen T.J., Jensen O.N., Briand C., and Peyrot V. 2001. Phosphorylation and oligomerization states of native pig brain HSP90 studied by mass spectrometry. Eur. J. Biochem. 268: 2402–2407.

Gatti A. and Traugh J.A. 1999. A two-dimensional peptide gel electrophoresis system for phosphopeptide mapping and amino acid sequencing. Anal. Biochem. 266: 198–204.

Gibson B.W. and Cohen P. 1990. Liquid secondary ion mass spectrometry of phosphorylated and sulfated peptides and proteins. Methods Enzymol. 193: 480–501.

Gobom J., Nordhoff E., Mirgorodskaya E., Ekman R., and Roepstorff P. 1999. Sample purification and preparation technique based on nano-scale reversed-phase columns for the sensitive analysis of complex peptide mixtures by matrix-assisted laser desorption/ionization mass spectrometry. J. Mass Spectrom. 34: 105–116.

Godovac-Zimmermann J. and Brown L.R. 2001. Perspectives for mass spectrometry and functional proteomics. Mass Spectrom. Rev. 20: 1–57.

Godovac-Zimmermann J., Soskic V., Poznanovic S., and Brianza F. 1999. Functional proteomics of signal transduction by membrane receptors. Electrophoresis 20: 952–961.

Gold M.R., Yungwirth T., Sutherland C.L., Ingham R.J., Vianzon D., Chiu R., Vanoostveen I., Morrison H.D., and Aebersold R. 1994. Purification and identification of tyrosine-phosphorylated proteins from b lymphocytes stimulated through the antigen receptor. Electrophoresis 15: 441–453.

Goodlett D.R., Aebersold R., and Watts J.D. 2000a. Quantitative in vitro kinase reaction as a guide for phosphoprotein analysis by mass spectrometry. Rapid Commun. Mass Spectrom. 14: 344–348.

Goodlett D.R., Bruce J.E., Anderson G.A., Rist B., Pasa-Tolic L., Fiehn O., Smith R.D., and Aebersold R. 2000b. Protein identification with a single accurate mass of a cysteine-containing peptide and constrained database searching. Anal. Chem. 72: 1112–1118.

Goodlett D.R., Wahl J.H., Udseth H.R., and Smith R.D. 1993. Reduced elution speed detection for capillary electrophoresis-mass spectrometry. J. Microcol. Sep. 5: 57–61.

Goshe M.B., Conrads T.P., Panisko E.A., Angell N.H., Veenstra T.D., and Smith R.D. 2001. Phosphoprotein isotope-coded affinity tag approach for isolating and quantitating phosphopeptides in proteome-wide analyses. Anal. Chem. 73: 2578–2586.

Gygi S.P., Rist B., Gerber S.A., Turecek F., Gelb M.H., and Aebersold R. 1999. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. Nat. Biotechnol. 17: 994–999.

Haebel S., Jensen C., Andersen S.O., and Roepstorff P. 1995. Isoforms of a cuticular protein from larvae of the meal beetle, Tenebrio molitor, studied by mass spectrometry in combination with Edman degradation and two-dimensional polyacrylamide gel electrophoresis. Protein Sci. 4: 394–404.

Hanger D.P., Betts J.C., Loviny T.L.F., Blackstock W.P., and Anderton B.H. 1998. New phosphorylation sites identified in hyperphosphorylated tau (paired helical filament-tau) from alzheimer's disease brain using nanoelectrospray mass spectrometry. J. Neurochem. 71: 2465–2476.

Hart S.R., Waterfield M.D., Burlingame A.L., and Cramer R. 2002. Factors governing the solubilization of phosphopeptides retained on ferric NTA IMAC beads and their analysis by MALDI TOFMS. J. Am. Soc. Mass Spectrom. 13: 1042–1051.

Hayashi F., Underhill D.M., Ozinsky A., Smith K.D., Yi E.C., Eng J.K., Goodlett D.R., and Aderem A. 2001. The innate immune response to bacterial flagellin is mediated by Toll-like receptor 5. Nature 410: 1099– 1103.

Huddleston M.J., Annan R.S., Bean M.F., and Carr S.A. 1993. Selective detection of phosphopeptides in complex-mixtures by electrospray liquid-chromatography mass-spectrometry. J. Am. Soc. Mass Spectrom. 4: 710–715.

Huddleston M.J., Annan R.S., Bean M.F., and Carr S.A. 1994. Selective detecton of Thr-, Ser-, and Tyr-phosphopeptides in complex digests by electrospray LC-MS. In Techniques in protein chemistry V (ed. J.W. Crabb), pp. 123–130. Academic Press, San Diego, California.

Hunter T. 1995. Protein kinases and phosphatases: The yin and yang of protein phosphorylation and signaling. Cell 80: 225–236.

^_______. 1998. The Croonian Lecture 1997. The phosphorylation of proteins on tyrosine: Its role in cell growth and disease. Phil. Trans. R. Soc. Lond. B 353: 583–605.

^_______. 2000. Signaling—2000 and beyond. Cell 100: 113–127.

Hunter A.P. and Games D.E. 1994. Chromatographic and mass spectrometric methods for the identification of phosphorylation sites in phosphoproteins. Rapid Commun. Mass Spectrom. 8: 559–565.

Immler D., Gremm D., Kirsch D., Spengler B., Presek P., and Meyer H.E. 1998. Identification of phosphorylated proteins from thrombin-activated human platelets isolated by two-dimensional gel electrophoresis by electrospray ionization-tandem mass spectrometry (ESI-MS/MS) and liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS). Electrophoresis 19: 1015–1023.

Jaffe H., Veeranna, and Pant H.C. 1998. Characterization of serine and threonine phosphorylation sites in beta-elimination/ethanethiol addition-modified proteins by electrospray tandem mass spectrometry and database searching. Biochemistry 37: 16211–16224.

Janek K., Wenschuh H., Bienert M., and Krause E. 2001. Phosphopeptide analysis by positive and negative ion matrix-assisted laser desorption/ionization. Rapid Commun. Mass Spectrom. 15: 1593–1599.

Jensen O.N. 2000. Modification-specific proteomics: Systematic strategies for analysing post-translationally modified proteins. In Proteomics: A trends guide (ed. W. Blackstock and M. Mann), pp. 36–42. Elsevier Science, Amsterdam.

Jensen O.N., Shevchenko A., and Mann M. 1997. Protein analysis by mass spectrometry. In Protein structure: A practical approach (ed. T.E. Creighton), pp. 29–57. IRL Press, Oxford, United Kingdom.

Jensen O.N., Wilm M., Shevchenko A., and Mann M. 1999. Peptide sequencing of 2-DE gel-isolated proteins by nanoelectrospray tandem mass spectrometry. Methods Mol. Biol. 112: 571–588.

Ji H., Baldwin G.S., Burgess A.W., Moritz R.L., Ward L.D., and Simpson R.J. 1993. Epidermal growth factor induces serine phosphorylation of stathmin in a human colon carcinoma cell line (LIM 1215). J. Biol. Chem. 268: 13396–13405.

Johnson L.N. and Lewis R.J. 2001. Structural basis for control by phosphorylation. Chem Rev. 101: 2209–2242.

Karas M. and Hillenkamp F. 1988. Laser desorption ionization of proteins with molecular masses exceeding 10,000 daltons. Anal. Chem. 60: 2299–2301.

Karlsson K.E. and Novotny M. 1988. Separation efficiency of slurry-packed liquid chromatography microcolumns with very small inner diameters. Anal. Chem. 60: 1662–1665.

Katta V., Chowdhury S.K., Chait B.T. 1991. Use of a single-quadrupole mass spectrometer for collision-induced dissociation studies of multiply charged peptide ions produced by electrospray ionization. Anal. Chem. 63: 174–179.

Katze M.G., Kwieciszewski B., Goodlett D.R., Blakely C.M., Nedderman P., Tan S.-L, and Aebersold R. 2000. Ser(2194) is a highly conserved major phosphorylation site of the hepatitis C virus nonstructural protein NS5A. Virology 278: 501–513.

Kelleher N.L., Zubarev R.A., Bush K., Furie B., Furie B.C., McLafferty F.W., and Walsh C.T. 1999. Localization of labile posttranslational modifications by electron capture dissociation: The case of gamma-carboxyglutamic acid. Anal. Chem. 71: 4250–4253.

Kennedy R.T. and Jorgenson J. W. 1989. Quantitative analysis of individual neurons by open tubular liquid chromatography with voltammetric detection. Anal. Chem. 61: 1128–1135.

Kolesnikova V.Y., Sklyankina V.A., Baratova L.A., Nazarova T.I., and Avaeva S.W. 1974. Modification of O-phosphoserine residues in phosphoproteins. Biochemistry 39: 235–240.

Krebs E.G. 1994. The growth of research on protein phosphorylation. Trends Biol. Sci. 19: 439–439.

Krishna R.G. and Wold F. 1993. Post-translational modification of proteins. Adv. Enzymol. Relat. Areas Mol. Biol. 67: 265–298.

Lapko V.N., Jiang X.Y., Smith D.L., and Song P.S. 1997. Posttranslational modification of oat phytochrome A: Phosphorylation of a specific serine in a multiple serine cluster. Biochemistry 36: 10595–10599.

Larsen M.R., Sørensen G.L., Fey S.J., Larsen P.M., and Roepstorff P. 2001. 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. Proteomics 1: 223–238.

Lee C.H., McComb M.E., Bromirski M., Jilkine A., Ens W., Standing K.G., and Perreault H. 2001. On-membrane digestion of beta-casein for determination of phosphorylation sites by matrix-assisted laser desorption/ionization quadrupole/time-of-flight mass spectrometry. Rapid Commun. Mass Spectrom. 15: 191–202.

Lehr S., Herkner A., Sickmann A., Meyer H.E., Krone W., and Müller-Wieland D. 2000. Identification of major tyrosine phosphorylation sites in the human insulin receptor substrate Gab-1 by insulin receptor kinase in vitro. Biochemistry 39: 10898–10907.

Li S. and Dass C. 1999. Iron(III)-immobilized metal ion affinity chromatography and mass spectrometry for the purification and characterization of synthetic phosphopeptides. Anal. Biochem. 270: 9–14.

Liao P.C., Leykam J., Andrews P.C., Cage D.A., and Allison J. 1994. An approach to locate phosphorylation sites in a phosphoprotein: Mass mapping by combining specific enzymatic degradation with matrix-assisted laser desorption/ionization mass spectrometry. Anal. Biochem. 219: 9–20.

Loboda A.V., Krutchinsky A.N., Bromirski M., Ens W., and Standing K.G. 2000. A tandem quadrupole/time-of-flight mass spectrometer with a matrix- assisted laser desorption/ionization source: Design and performance. Rapid Commun. Mass Spectrom. 14: 1047–1057.

Ma Y., Lu Y., Mo W., and Neubert T.A. 2001. Rapid detection of phosphopeptides from protein digests using matrix assisted laser desorption/ionization time-of-flight mass spectrometry and nanoelectrospray quadrupole time-of-flight mass spectrometry. Rapid Commun. Mass Spectrom. 15: 1693–1700.

MacDonald J.A., Mackey A.J., Pearson W.R., and Haystead T.A.J. 2002. A strategy for the rapid identification of phosphorylation sites in the phoshoproteome. Mol. Cell. Proteomics 1: 314–322.

Mann M., Hendrickson R.C., and Pandey A. 2001. Analysis of proteins and proteomes by mass spectrometry. Annu. Rev. Biochem. 70: 437–473.

Mann M., Ong S.E., Gronborg M., Steen H., Jensen O.N., and Pandey A. 2002. Analysis of protein phosphorylation using mass spectrometry: Deciphering the phosphoproteome. Trends Biotechnol. 20: 261–268.

Marcus K., Immler D., Sternberger J., and Meyer H.E. 2000. Identification of platelet proteins separated by two-dimensional gel electrophoresis and analyzed by matrix assisted laser desorption/ionization-time of flight-mass spectrometry and detection of tyrosine-phosphorylated proteins. Electrophoresis 21: 2622–2636.

Merrick B.A., Zhou W., Martin K.J., Jeyarajah S., Parker C.E., Selkirk J.K., Tomer K.B., and Borchers C.H. 2001. Site-specific phosphorylation of human p53 protein determined by mass spectrometry. Biochemistry 40: 4053–4066.

Martensen T.M. 1971. Chemical properties, isolation, and analysis of O-phosphates in proteins. Methods Enzymol. 107: 3–23.

Meyer H., Hoffmann-Posorske E., Korte H., and Heilmeyer L.J. 1986. Sequence analysis of phosphoserine-containing peptides. Modification for picomolar sensitivity. FEBS Lett. 204: 61–66.

Meyer H.E., Eisermann B., Heber M., Hoffmann-Posorske E., Korte H., Weigt C., Wegner A., Hutton T., Donella-Deana A., and Perich J.W. 1993. Strategies for nonradioactive methods in the localization of phosphorylated amino acids in proteins. FASEB J. 7: 776–782.

Michel H., Hunt D.F., Shabanowitz J., and Bennett J. 1988. Tandem mass spectrometry reveals that three photosystem II proteins of spinach chloroplasts contain N-acetyl-O-phosphothreonine at their NH₂ termini. J. Biol. Chem. 263: 1123–1130.

Miliotis T., Ericsson P.O., Marko-Varga G., Svensson R., Nilsson J., Laurell T., and Bischoff R. 2001. Analysis of regulatory phosphorylation sites in ZAP-70 by capillary high-performance liquid chromatography coupled to electrospray ionization or matrix-assisted laser desorption ionization time-of- flight mass spectrometry. J. Chromatogr. B 752: 323–334.

Muszynska G., Dobrowolska G., Medin A., Ekamn P., and Porath J.O. 1992. Model studies on iron (III) ion affinity chromatography II. Interaction of immobilized iron (III) ions with phosphorylated amino acids, peptides and proteins. J. Chromatogr. 604: 19–28.

Neubauer G. and Mann M. 1999. Mapping of phosphorylation sites of gel-isolated proteins by nanoelectrospray tandem mass spectrometry: Potentials and limitations. Anal. Chem. 71: 235–242.

Neville D.C.A., Rozanas C.R., Price E.M., Gruis D.B., Verkman A.S., and Townsend R.R. 1997. Evidence for phosphorylation of serine 753 in CFTR using a novel metal-ion affinity resin and matrix-assisted laser desorption mass spectrometry. Protein Sci. 6: 2436–2445.

Nuwaysir L.M. and Stults J.T. 1993. electrospray ionization mass spectrometry of phosphopeptides isolated by on-line immobilized metal-ion affinity chromatography. J. Am. Soc. Mass Spectrom. 4: 662–669.

Oda Y., Nagasu T., and Chait B.T. 2001. Enrichment analysis of phosphorylated proteins as a tool for probing the phosphoproteome. Nat. Biotechnol. 19: 379–382.

Oda Y., Huang K., Cross F.R., Cowburn D., and Chait B.T. 1999. Accurate quantitation of protein expression and site-specific phosphorylation. Proc. Natl. Acad. Sci. 96: 6591–6596.

Olcott M.C., Bradley M.L., and Haley B.E. 1994. Photoaffinity labeling of creatine kinase with 2-azido- and 8-azidoadenosine triphosphate: Identification of two peptides from the ATP-binding domain. Biochem. 33: 11935–11941.

Pandey A., Andersen J.S., and Mann M. 2000a. Use of mass spectrometry to study signaling pathways. Sci. STKE 37: PL1.

Pandey A., Podtelejnikov A.V., Blagoev B., Bustelo X.R., Mann M., and Lodish H.F. 2000b. Analysis of receptor signaling pathways by mass spectrometry: Identification of Vav-2 as a substrate of the epidermal and platelet-derived growth factor receptors. Proc. Natl. Acad. Sci. 97: 179–184.

Papac D.I., Hoyes J., and Tomer K.B. 1994. Direct Analysis of Affinity-Bound Analytes by MALDI/TOF MS. Anal. Chem. 66: 2609–2613.

Pas H.H. and Robillard G.T. 1988. S-phosphocysteine and phosphohistidine are intermediates in the phosphoenolpyruvate-dependent mannitol transport catalyzed by Escherichia coli EIIMtl. Biochemistry 27: 5835–5839.

Porath J., Carlsson J., Olsson I., and Belfrage G. 1975. Metal chelate affinity chromatography, a new approach to protein fractionation. Nature 258: 598–599.

Posada J. and Cooper J.A. 1992. Molecular signal integration. Interplay between serine, threonine, and tyrosine phosphorylation. Mol. Biol. Cell 3: 583–592.

Posewitz M.C. and Tempst P. 1999. Immobilized gallium(III) affinity chromatography of phosphopeptides. Anal. Chem. 71: 2883–2892.

Powell K.A., Valova V.A., Malladi C.S., Jensen O.N., Larsen M.R., and Robinson P.J. 2000. Phosphorylation of dynamin I on Ser-795 by protein kinase C blocks its association with phospholipids. J. Biol. Chem. 275: 11610–11617.

Qian X.-H., Zhou W., Khaledi M.G., and Tomer K.B. 1999. Direct analysis of the products of sequential cleavages of peptides and proteins affinity-bound to immobilized metal ion beads by matrix-assisted laser desorption/ionization mass spectrometry. Anal. Biochem. 274: 174–180.

Qin J. and Chait B.T. 1997. Identification and characterization of posttranslational modifications of proteins by MALDI ion trap mass spectrometry. Anal. Chem. 69: 4002–4009.

Qin J., Fenyo D., Zhao Y.M., Hall W.W., Chao D.M., Wilson C.J., Young R.A., and Chait B.T. 1997. A strategy for rapid, high confidence protein identification. Anal. Chem. 69: 3995–4001.

Raska C.S., Parker C.E., Dominski Z., Marzluff W.F., Glish G.L., Pope R.M., and Borchers C.H. 2002. Direct MALDI-MS/MS of phosphopeptides affinity-bound to immobilized metal ion affinity chromatography beads. Anal. Chem. 74: 3429–3433.

Resing K.A., Johnson R.S., and Walsh K.A. 1995. Mass spectrometric analysis of 21 phosphorylation sites in the internal repeat of rat profilaggrin, precursor of an intermediate filament associated protein. Biochemistry 34: 9477–9487.

Roepstorff P. and Fohlmann J. 1984. Proposal for a common nomenclature for sequence ions in mass spectra of peptides. Biomed. Mass Spectrom. 11: 601.

Schlessinger J. 2000. Cell signaling by receptor tyrosine kinases. Cell 103: 211–225.

Schlosser A., Pipkorn R., Bossemeyer D., and Lehmann W.D. 2000. Analyse der Proteinphosphorylierung durch Kombination von Elastase-Verdau und ESI-Tandem-Massenspektrometrie. In Proceedings of the 32nd annual meeting of the German Mass Spectrometry Society, p. 115. German Mass Spectrometry Society, Oldenburg, Germany.

^_______. 2001. Analysis of protein phosphorylation by a combination of elastase digestion and neutral loss tandem mass spectrometry. Anal. Chem. 73: 170–176.

Schrecker O., Stein R., Hengstenberg W., Gassner M., and Stehlik D. 1975. The staphylococcal PEP dependent phosphotransferase system, proton magnetic resonance (PMR) studies on the phosphoryl carrier protein HPr: Evidence for a phosphohistidine residue in the intact phospho-HPr molecule. FEBS Lett. 51: 309–312.

Shevchenko A., Loboda A., Ens W., and Standing K.G. 2000. MALDI quadrupole time-of-flight mass spectrometry: A powerful tool for proteomic research. Anal. Chem. 72: 2132–2141.

Shevchenko A., Wilm M., Vorm O., and Mann M. 1996. Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal. Chem. 68: 850–858.

Shi S.D., Hemling M.E., Carr S.A., Horn D.M., Lindh I., and McLafferty F.W. 2001. Phosphopeptide/phosphoprotein mapping by electron capture dissociation mass spectrometry. Anal. Chem. 73: 19–22.

Sickmann A. and Meyer H.E. 2001. Phosphoamino acid analysis. Proteomics 1: 200–206.

Sickmann A., Dormeyer W., Wortelkamp S., Woitalla D., Kuhn W., and Meyer H.E. 2000. Identification of proteins from human cerebrospinal fluid, separated by two-dimensional polyacrylamide gel electrophoresis. Electrophoresis 21: 2721–2728.

Springer M.S., Coy M.F., and Adler J. 1979. Protein methylation in behavioural control mechanisms and in signal transduction. Nature 280: 279–284.

Steen H. and Mann M. 2002. A new derivatization strategy for the analysis of phosphopeptides by precursor ion scanning in positive ion mode. J. Am. Soc. Mass Spectrom. 13: 996–1003.

Steen H., Küster B., and Mann M. 2001a. Quadrupole time-of-flight versus triple-quadrupole mass spectrometry for the determination of phosphopeptides by precursor ion scanning. J. Mass Spectrom. 36: 782–790.

Steen H., Küster B., Fernandez M., Pandey A., and Mann M. 2001b. Detection of tyrosine phosphorylated peptides by precursor ion scanning quadrupole TOF mass spectrometry in positive ion mode. Anal. Chem. 73: 1440–1448.

Stensballe A., Andersen S., and Jensen O.N. 2001. Characterization of phosphoproteins from electrophoretic gels by nanoscale Fe(III) affinity chromatography with off-line mass spectrometry analysis. Proteomics 1: 207–222.

Stensballe A., Jensen O.N., Olsen J.V., Haselmann K.F., and Zubarev R.A. 2000. Electron capture dissociation of singly and multiply phosphorylated peptides. Rapid Commun. Mass Spectrom. 14: 1793–1800.

Storm S.M. and Khawaja X.Z. 1999. Probing for drug-induced multiplex signal transduction pathways using high resolution two-dimensional gel electrophoresis: Application to beta-adrenoceptor stimulation in the rat C6 glioma cell. Brain. Res. Mol. Brain. Res. 71: 50–60.

Vihinen H. and Saarinen J. 2000. Phosphorylation site analysis of semliki forest virus nonstructural protein 3. J. Biol. Chem. 275: 27775–27783.

Wahl J.H., Goodlett D.R., Udseth H.R., and Smith R.D. 1993. Use of small-diameter capillaries for increasing peptide and protein detection sensitivity in capillary electrophoresis-mass spectrometry. Electrophoresis 14: 448–457.

Weckwerth W., Willmitzer L., and Fiehn O. 2000. Comparative quantification and identification of phosphoproteins using stable isotope labeling and liquid chromatography/mass spectrometry. Rapid Commun. Mass Spectrom. 14: 1677–1681.

Weijland A., Williams J.C., Neubauer G., Courtneidge S.A., Wierenga R.K., and Superti-Furga G. 1997. Src regulated by C-terminal phosphorylation is monomeric. Proc. Natl. Acad. Sci. 94: 3590–3595.

Wettenhall R.E.H., Aebersold R., and Hood L.E. 1991. Solid-Phase Sequencing of ³²p-labeled phosphopeptides at picomole and subpicomole levels. Methods Enzymol. 201: 186–199.

Wilm M., Neubauer G., and Mann M. 1996. Parent ion scans of unseparated peptide mixtures. Anal. Chem. 68: 527–533.

Wind M., Edler M., Jakubowski N., Linscheid M., Wesch H., and Lehmann W.D. 2001. Analysis of protein phosphorylation by capillary liquid chromatography coupled to element mass spectrometry with 31P detection and to electrospray mass spectrometry. Anal. Chem. 73: 29–35.

Yan J.X., Packer N.H., Gooley A.A., and Williams K.L. 1998. Protein phosphorylation: Technology for the identification of phosphoamino acids. J. Chromatogr. 808: 23–41.

Zhang X.L., Herring C.J., Romano P.R., Szczepanowska J., Brzeska H., Hinnebusch A.G., and Qin J. 1998. Identification of phosphorylation sites in proteins separated by polyacrylamide gel electrophoresis. Anal. Chem. 70: 2050–2059.

Zhao J.Y., Kuang J., Adlakha R.C., and Rao P.N. 1989. Threonine phosphorylation is associated with mitosis in HeLa cells. FEBS Lett. 249: 389–395.

Zhou H., Watts J.D., and Aebersold R. 2001. A systematic approach to the analysis of protein phosphorylation. Nat. Biotechnol. 19: 375–378.

Zhou W., Merrick B.A., Khaledi M.G., and Tomer K.B. 2000. Detection and sequencing of phosphopeptides affinity bound to immobilized metal ion beads by matrix-assisted laser desorption/ionization mass spectrometry. J. Am. Soc. Mass Spectrom. 11: 273–282.

Zubarev R.A., Kelleher N.L., and McLafferty F.W. 1998. Electron capture dissociation of multiply charged protein cations. A nonergodic process. J. Am. Chem. Soc. 120: 3265–3266.