Development of high-throughput workflow for glycosphingolipid analysis and annotation
开发用于糖脂分析和注释的高通量工作流程
基本信息
- 批准号:9167031
- 负责人:
- 金额:$ 32.74万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2016
- 资助国家:美国
- 起止时间:2016-08-10 至 2018-07-31
- 项目状态:已结题
- 来源:
- 关键词:AlgorithmsAnabolismAutomated AnnotationBiocompatible MaterialsBiologicalBiological ProcessCellsCeramidesCommunitiesComputer AssistedComputer SimulationComputer softwareCoupledDataData AnalysesData SetDatabasesDevelopmentDigestionDiseaseDocumentationEducationEnzymesEukaryotic CellEventGangliosidosis GM1GenomicsGlycolipidsGlycosphingolipidsGoalsGrantGrowthHarvestHeterogeneityHumanLeadLibrariesLipidsLiquid ChromatographyMainstreamingManualsMass Spectrum AnalysisMediatingMembraneMethodologyMethodsMusPlayPolysaccharidesPreparationProteomicsProtocols documentationResearch PersonnelResourcesRoleSamplingSignal TransductionSoftware ToolsSourceStructureTeaching MaterialsTechniquesTimebasebrain tissuecomputerized data processingdesignenzyme substrateinstrumentinstrumentationliquid chromatography mass spectrometrymeetingspathogenpreferencereceptorsoftware developmenttoolweb site
项目摘要
PROJECT SUMMARY
Glycosphingolipids (GSL) are essential components of all biological membranes and serve as receptors and
modulators of cell-cell and cell-pathogen interactions as well as regulators of diverse receptor-mediated cell
signaling events. Highly sensitive, rapid, and facile methods for analyzing GSL structures harvested from
biological sources do not currently exist, but would catalyze broader participation by non-glycoscientists in
understanding GSL function. Most current glycomics approaches for the characterization of GSL structure
utilize enzymatic digestion to release the hydrophilic glycan from the hydrophobic lipid. While this approach
simplifies the complexity of the analyte, it blinds the analyst from detecting lipid heterogeneity and, thereby,
ignores the biological importance of the lipid portion of the GSLs. Furthermore, enzyme-substrate preferences
bias glycan profiles generated by digestion. Liquid chromatography (LC) of released GSL glycans coupled to
mass spectrometry (MS) have been broadly applied to quantifying and characterizing GSL profiles.
Unfortunately, LC-MS based analytic approaches are not able to profile both neutral and acidic GSL
components at the same time and these workflows generally require time-consuming sample preparation and
intensive manual interpretation of the data. To remedy the consequences of enzyme bias and chromatographic
complexity, we will develop, a sensitive, robust and comprehensive methodology for glycosphingolipidomics
supported by a semi-automated annotation software with a highly-curated database to further understand the
biological significances of GSLs.
We propose to develop standardized methods for the preparation and identification of intact GSL without
using enzymatic release of the glycan from the lipid portion. Furthermore, these methods will employ
permethylation to enhance structural analysis and to support the simultaneous identification of neutral and
acidic GSL structures. As part of the proposed workflow we will optimize this method for different MS analytical
platforms and fragmentation types. The sample preparation and MS methods will be complimented by a
software suite that facilitates analysis and annotation of the resulting datasets. The software will support high
throughput MS data analysis by significantly reducing the time required for annotation and interpretation.
Sample preparation protocols as well as MS methods for different platforms and fragmentation types will be
made publicly available to the community on our project website. The software will be freely available for
download as well.
项目摘要
鞘糖脂(GSL)是所有生物膜的必需组分,并且作为受体和
细胞-细胞和细胞-病原体相互作用的调节剂以及多种受体介导的细胞
信号事件。高度灵敏、快速和简便的方法,用于分析从
生物来源目前尚不存在,但将促进非糖科学家更广泛地参与
了解GSL功能。用于GSL结构表征的最新糖组学方法
利用酶消化从所述疏水性脂质释放所述亲水性聚糖。虽然这种方法
简化了分析物的复杂性,使分析者无法检测脂质异质性,因此,
忽略了GSL脂质部分的生物学重要性。此外,酶-底物偏好
通过消化产生的偏向聚糖谱。释放的GSL聚糖偶联至
质谱(MS)已被广泛应用于定量和表征GSL谱。
不幸的是,基于LC-MS的分析方法不能分析中性和酸性GSL
这些工作流程通常需要耗时的样品制备,
对数据进行密集的人工解释。为了弥补酶偏差和色谱分析的后果,
复杂性,我们将开发一种敏感,稳健和全面的鞘糖脂组学方法
由一个半自动化的注释软件与高度策划的数据库支持,以进一步了解
GSL的生物学意义。
我们建议开发标准化的方法,用于制备和鉴定完整的GSL,
使用聚糖从脂质部分的酶促释放。此外,这些方法将采用
全甲基化以增强结构分析并支持同时鉴定中性和
酸性GSL结构。作为拟定工作流程的一部分,我们将针对不同的MS分析优化该方法。
平台和碎片化类型。样品制备和MS方法将通过
软件套件,便于分析和注释的结果数据集。该软件将支持高
通过显著减少注释和解释所需的时间,提高了MS数据分析的吞吐量。
将介绍不同平台和碎片类型的样品制备方案以及MS方法。
在我们的项目网站上向社区公开。该软件将免费提供给
下载也是。
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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Kazuhiro Aoki其他文献
Kazuhiro Aoki的其他文献
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{{ truncateString('Kazuhiro Aoki', 18)}}的其他基金
Development of high-throughput workflow for glycosphingolipid analysis and annotation
开发用于糖脂分析和注释的高通量工作流程
- 批准号:
9330073 - 财政年份:2016
- 资助金额:
$ 32.74万 - 项目类别:
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