Zhang Wei, Cao Weiqian, Huang Jiangming, Wang Hong, Wang Ji, Xie Chen, Yang Pengyuan
Department of Chemistry, Fudan University, Shanghai 200433, P. R. China.
Analyst. 2015 Feb 21;140(4):1082-9. doi: 10.1039/c4an02073a.
PNGase F-catalyzed glycosylation site (18)O-labeling is a widely used method for glycoprotein quantitation owing to its efficiency and simplicity. However, PNGase F-catalyzed glycan (18)O-labeling, which offers advantages for glycomics, has not yet been developed. In this study, PNGase F-mediated incorporation of (18)O into glycans during the N-glycan release from glycoproteins by PNGase F was finally realized, named as PCGOL (PNGase F-catalyzed glycan (18)O-labeling), which offers a potential strategy for relative glycan quantitation. This new method showed good linearity and high reproducibility within at least 2 orders of magnitude in the dynamic range. Furthermore, PCGOL combined with our previously developed TOSIL method (tandem (18)O stable isotope labeling for N-glycoproteome quantitation) can be used for comprehensive N-glycosylation quantification, achieving simultaneous quantification of glycans, glycopeptides and glycoproteins in a single workflow, which was also used to analyze glycosylation changes in immunoglobulin G (IgG) associated with hepatocellular carcinoma in the present work.
PNGase F催化的糖基化位点（18）O标记因其高效性和简便性，是一种广泛用于糖蛋白定量的方法。然而，PNGase F催化的聚糖（18）O标记虽对糖组学具有优势，但尚未得到开发。在本研究中，最终实现了PNGase F介导的在PNGase F从糖蛋白释放N-聚糖过程中将（18）O掺入聚糖，命名为PCGOL（PNGase F催化的聚糖（18）O标记），它为相对聚糖定量提供了一种潜在策略。这种新方法在动态范围内至少2个数量级内显示出良好的线性和高重现性。此外，PCGOL与我们先前开发的TOSIL方法（用于N-糖蛋白组定量的串联（18）O稳定同位素标记）相结合，可用于全面的N-糖基化定量，在单个工作流程中实现聚糖、糖肽和糖蛋白的同时定量，在本研究中还用于分析与肝细胞癌相关的免疫球蛋白G（IgG）的糖基化变化。
