Glycoproteomic Sample Processing, LC-MS, and Data Analysis Using GlycReSoft.
Glycoproteomic Sample Processing, LC-MS, and Data Analysis Using GlycReSoft.
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糖蛋白质组学样本处理、液相色谱 - 质谱联用（LC - MS）以及使用GlycReSoft进行数据分析
DOI：
10.1002/cpz1.84
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发表时间：
2021-03
期刊：
影响因子：
--
通讯作者：
Zaia J
中科院分区：
文献类型：
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作者：
Wang M;Shajahan A;Pepi LE;Azadi P;Zaia J
鉴定蛋白质特定位点上的N -糖基化和O -糖基化以及聚糖结构信息，对于确定糖蛋白在正常和病理细胞功能中所起的作用是必要的。由于这种糖基化存在宏观和微观的不均一性，并改变糖肽的解离行为，因此需要特定的样品制备、质谱分析和数据处理技术。基于先进串联质谱的糖蛋白质组学，结合强大的数据挖掘算法，是表征蛋白质糖基化的关键要素。本实验方案包括详细、简化的样品制备方法，用于液相色谱 - 质谱数据采集，以及随后使用公开可用的GlycReSoft程序进行基于生物信息学的数据注释，以高效鉴定和定量糖蛋白糖基化。
纯化糖蛋白上聚糖的表征及位点占有率
糖蛋白的胶内酶解
通过糖肽富集检测细胞/组织中的糖蛋白
通过高分辨率纳流液相色谱 - 串联质谱获取糖肽
使用GlycReSoft鉴定和定量糖肽
Identification of N- and O-glycosylation on specific sites of proteins, along with the glycan structural information, is necessary to determine the roles glycoproteins play in normal and pathological cellular functions. Because such glycosylation is macro and micro-heterogeneous and alters the dissociation behavior of glycopeptides, specific sample preparation, mass spectrometry, and data analysis techniques are required. Advanced tandem mass spectrometry based glycoproteomics coupled with powerful data mining algorithms are key elements for characterization of protein glycosylation. This protocol includes the detailed, streamlined sample preparation method for liquid chromatography-mass spectrometry data acquisition and subsequent bioinformatics-based data annotation using the publically available GlycReSoft program for highly efficient identification and quantification of glycoprotein glycosylation. CHARACTERIZATION OF GLYCANS AND THE SITE OCCUPANCY ON PURIFIED GLYCOPROTEIN IN-GEL DIGESTION OF GLYCOPROTEINS DETECTION OF GLYCOPROTEINS FROM CELLS/TISSUES THROUGH GLYCOPEPTIDE ENRICHMENT ACQUISITION OF GLYCOPEPTIDES THROUGH HIGH-RESOLUTION NANO-LC-MS/MS IDENTIFICATION AND QUANTIFICATION OF GLYCOPEPTIDES USING GlycReSoft