TiO(2) Simultaneous Enrichment, On-Line Deglycosylation, and Sequential Analysis of Glyco- and Phosphopeptides.
TiO(2) Simultaneous Enrichment, On-Line Deglycosylation, and Sequential Analysis of Glyco- and Phosphopeptides.
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TiO(2) 糖肽和磷酸肽的同步富集、在线去糖基化和顺序分析。
DOI：
10.3389/fchem.2021.703176
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发表时间：
2021
影响因子：
5.5
通讯作者：
Liang X
中科院分区：
文献类型：
--
作者：
Chen C;Zhang X;Dong X;Zhou H;Li X;Liang X
可逆的蛋白质糖基化和磷酸化紧密调节重要的细胞过程，并以串扰依赖性方式密切参与病理过程。由于糖肽和磷酸肽的重要性和低丰度，已经开发了几种策略来同时富集和共洗脱糖肽和磷酸肽。然而，去糖基化肽和磷酸肽的共存加剧了质谱分析。在此，我们开发了一种分析糖肽和磷酸肽的新策略，该策略基于同时富集 TiO2、在线去糖基化和收集去糖基化肽，以及随后洗脱磷酸肽。为了优化在线去糖基化条件，研究了溶液 pH、缓冲液类型和浓度以及去糖基化时间。将这种新策略应用到 100 μg 小鼠大脑中，产生了 355 种糖肽和 1,975 种磷酸肽，分别是用报道方法富集的 2.5 倍和 1.4 倍。这项研究将扩大 TiO2 的应用，并可能为同时监测蛋白质多重翻译后修饰提供线索。
Reversible protein glycosylation and phosphorylation tightly modulate important cellular processes and are closely involved in pathological processes in a crosstalk dependent manner. Because of their significance and low abundances of glyco- and phosphopeptides, several strategies have been developed to simultaneously enrich and co-elute glyco- and phosphopeptides. However, the co-existence of deglycosylated peptides and phosphopeptides aggravates the mass spectrometry analysis. Herein we developed a novel strategy to analyze glyco- and phosphopeptides based on simultaneous enrichment with TiO2, on-line deglycosylation and collection of deglycosylated peptides, and subsequent elution of phosphopeptides. To optimize on-line deglycosylation conditions, the solution pH, buffer types and concentrations, and deglycosylation time were investigated. The application of this novel strategy to 100 μg mouse brain resulted in 355 glycopeptides and 1,975 phosphopeptides, which were 2.5 and 1.4 folds of those enriched with the reported method. This study will expand the application of TiO2 and may shed light on simultaneously monitoring protein multiple post-translational modifications.
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