Enabling Mass Spectrometry Analysis of the Sulfoproteome
实现磺化蛋白质组的质谱分析
基本信息
- 批准号:10543467
- 负责人:
- 金额:$ 29.92万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2021
- 资助国家:美国
- 起止时间:2021-01-01 至 2024-11-30
- 项目状态:已结题
- 来源:
- 关键词:AcidityAlkanesulfonatesAmino AcidsAnionsArginineBasic Amino AcidsCationsCell Culture TechniquesCell divisionCell physiologyCell surfaceCellsChargeChloratesCollaborationsComplicationDataData AnalysesData SetDatabasesDetectionDevelopmentDiseaseDissociationElectron TransportElectronsElectrospray IonizationEnvironmentEnzymesEukaryotic CellFree RadicalsGORASP2 geneGasesGolgi ApparatusHIVHealthHeparinInfectionInflammationInterphaseIonsKnock-outLaboratoriesLiverLocationLysineManualsMass Spectrum AnalysisMeasurementMeasuresMembrane ProteinsMethodsMitoticModalityNatureNormal CellOrganellesPeptide Sequence DeterminationPeptidesPerformancePhasePhosphopeptidesPhosphoric Monoester HydrolasesPhosphorylationPlayPositioning AttributePost Translational Modification AnalysisPost-Translational Protein ProcessingProcessProtein RegionProtein SecretionProteinsRattusRegulationReportingRetinaRoleSiteSulfateSuraminTechniquesTechnologyTissuesTrifluoroethanolTyrosineValidationVesicleViralVirus DiseasesVirus ReplicationWorkabsorptionadductanimal tissuechemical propertydata acquisitiondetection methodexperimental studyextracellularfunctional groupglycosylationhepatoma cellimprovedinhibitorinstrumentinterestionizationknock-downliquid chromatography mass spectrometrymass spectrometermulti-photonnovelnovel strategiespeptide hormonephosphoproteomicspreventprotein protein interactionprotein-tyrosine sulfotransferasereceptor bindingrottlerinsulfotransferasesulfurtransferasetandem mass spectrometrytooltyrosine O-sulfate
项目摘要
Project Summary
Tyrosine O-sulfation, i.e., transfer of a sulfonate group to tyrosine amino acid residues in proteins, is a
widespread posttranslational modification (PTM) in eukaryotic cells with a variety of known functions in health
and disease, including receptor binding, viral replication, inflammation, and retinal function. The enzymes that
catalyze tyrosine sulfation are located in the Golgi apparatus. The function of this organelle is to ensure that
correct protein modification occurs and to package proteins into vesicles for export to the cell surface, or the
extracellular environment. Because proteins must typically enter the Golgi to become sulfated, most known
sulfoproteins are secreted proteins or membrane proteins. Mass spectrometry (MS) is a powerful tool for global
PTM analysis in cells and tissues; however, large scale analysis of tyrosine O-sulfation has not been feasible,
due in part to its labile nature in the gas-phase environment of a mass spectrometer, and in part due to the lack
of appropriate data analysis strategies. In MS experiments, proteins are typically digested into smaller peptides,
which are ionized, detected, and fragmented to deduce sequence information. When measuring protein
phosphorylation, another rather labile PTM known to regulate Golgi disassembly and reassembly during cell
division, in interphase vs. mitotic Golgi, we found that tyrosine O-sulfation was co-enriched. This discovery is
not surprising because the chemical properties of sulfation (O-SO3) are similar to phosphorylation (O-PO3H).
However; high mass accuracy measurements are required to deduce the small mass difference of 0.0095 Da
between these two PTMs. Even as such high performance measurements are becoming more routine, standard
database search tools typically do not identify protein sulfation because this PTM is completely lost during
analysis. We found that open database searching was able to overcome this problem and, thus, we were able
to accomplish identification of a number of novel sulfoproteins in rat liver Golgi. While an exciting advance, the
exact location of O-sulfation within proteolytic peptides could not be directly measured. In Aim 1 of this proposal,
we seek to develop improved methods for detection of intact sulfopeptides by MS, including elimination of
competing phosphorylation, determination of peptide sequence effects, implementation of stabilizing adducts,
and conditions that selectively dissociate sulfopeptides. To further allow sulfate site localization, in Aim 2, we
seek to develop technologies for fragmenting sulfopeptides while retaining sulfate in fragment ions. These
approaches include negative ion mode free radical initiated peptide sequencing, which allows sulfopeptides to
enter the mass spectrometer as more stable anions, and the development of “smart” data acquisition strategies
for improved electron transfer dissociation. The final Aim 3 seeks to apply these improved approaches for
comprehensive analysis of the Golgi sulfoproteome in cells and animal tissue, particularly under perturbed Golgi
conditions, which are expected to alter sulfation. These types of measurements will provide transformative
information regarding the regulatory roles of tyrosine sulfation and its impact on cellular function.
项目总结
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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KRISTINA HAKANSSON其他文献
KRISTINA HAKANSSON的其他文献
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{{ truncateString('KRISTINA HAKANSSON', 18)}}的其他基金
Enabling Mass Spectrometry Analysis of the Sulfoproteome
实现磺化蛋白质组的质谱分析
- 批准号:
10322358 - 财政年份:2021
- 资助金额:
$ 29.92万 - 项目类别:
Enabling Mass Spectrometry Analysis of the Sulfoproteome
实现磺化蛋白质组的质谱分析
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10096628 - 财政年份:2021
- 资助金额:
$ 29.92万 - 项目类别:
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- 资助金额:
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癌症干细胞聚糖结构测定的新方法
- 批准号:
8042696 - 财政年份:2010
- 资助金额:
$ 29.92万 - 项目类别:
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癌症干细胞聚糖结构测定的新方法
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- 资助金额:
$ 29.92万 - 项目类别:
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- 资助金额:
$ 29.92万 - 项目类别:














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