Qualitative and Quantitative Analyses of Protein Phosphorylation in Naive and Stimulated Mouse Synaptosomal Preparations

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dc.contributor.author Munton, RP en
dc.contributor.author Tweedie-Cullen, Ry en
dc.contributor.author Livingstone-Zatchej, M en
dc.contributor.author Weinandy, F en
dc.contributor.author Waidelich, M en
dc.contributor.author Longo, D en
dc.contributor.author Gehrig, P en
dc.contributor.author Potthast, F en
dc.contributor.author Rutishauser, D en
dc.contributor.author Gerrits, B en
dc.contributor.author Panse, C en
dc.contributor.author Schlapbach, R en
dc.contributor.author Mansuy, IM en
dc.date.accessioned 2013-10-09T20:59:12Z en
dc.date.issued 2007 en
dc.identifier.citation Molecular and Cellular Proteomics 6(2):283-293 2007 en
dc.identifier.issn 1535-9476 en
dc.identifier.uri http://hdl.handle.net/2292/20893 en
dc.description.abstract Activity-dependent protein phosphorylation is a highly dynamic yet tightly regulated process essential for cellular signaling. Although recognized as critical for neuronal functions, the extent and stoichiometry of phosphorylation in brain cells remain undetermined. In this study, we resolved activity-dependent changes in phosphorylation stoichiometry at specific sites in distinct subcellular compartments of brain cells. Following highly sensitive phosphopeptide enrichment using immobilized metal affinity chromatography and mass spectrometry, we isolated and identified 974 unique phosphorylation sites on 499 proteins, many of which are novel. To further explore the significance of specific phosphorylation sites, we used isobaric peptide labels and determined the absolute quantity of both phosphorylated and non-phosphorylated peptides of candidate phosphoproteins and estimated phosphorylation stoichiometry. The analyses of phosphorylation dynamics using differentially stimulated synaptic terminal preparations revealed activity-dependent changes in phosphorylation stoichiometry of target proteins. Using this method, we were able to differentiate between distinct isoforms of Ca2+/calmodulin-dependent protein kinase (CaMKII) and identify a novel activity-regulated phosphorylation site on the glutamate receptor subunit GluR1. Together these data illustrate that mass spectrometry-based methods can be used to determine activity-dependent changes in phosphorylation stoichiometry on candidate phosphopeptides following large scale phosphoproteome analysis of brain tissue. en
dc.language Eng en
dc.publisher The American Society for Biochemistry and Molecular Biology en
dc.relation.ispartofseries Molecular and Cellular Proteomics en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. Details obtained from http://www.jbc.org/site/misc/Copyright_Permission.xhtml en
dc.rights.uri https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm en
dc.title Qualitative and Quantitative Analyses of Protein Phosphorylation in Naive and Stimulated Mouse Synaptosomal Preparations en
dc.type Journal Article en
dc.identifier.doi 10.1074/mcp.M600046-MCP200 en
pubs.issue 2 en
pubs.begin-page 283 en
pubs.volume 6 en
dc.identifier.pmid 17114649 en
pubs.end-page 293 en
dc.rights.accessrights http://purl.org/eprint/accessRights/RestrictedAccess en
pubs.subtype Article en
pubs.elements-id 407112 en
pubs.org-id Medical and Health Sciences en
pubs.org-id School of Medicine en
pubs.org-id Medicine Department en
dc.identifier.eissn 1535-9484 en
pubs.record-created-at-source-date 2013-10-10 en
pubs.dimensions-id 17114649 en


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