Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433, China.
Anal Chem. 2011 Jun 15;83(12):4975-81. doi: 10.1021/ac200753e. Epub 2011 May 27.
Stable isotopic labeling coupled with mass spectrometry analysis is a promising method of detecting quantitative variations in glycans, which may result in aberrant glycosylation in many disorders and diseases. Although various isotopic labeling methods have been used for relative glycan quantitation, enzymatic (18)O labeling, which offers advantages for glycomics similar to those by protease-catalyzed (18)O labeling for proteomics, has not been developed yet. In this study, endoglycosidase incorporated (18)O into the N-glycan reducing end in (18)O-water as N-glycans were released from glycoproteins, rendering glycan reducing-end (18)O labeling (GREOL) a potential strategy for relative glycan quantitation. This proposed method provided good linearity with high reproducibility within 2 orders of magnitude in dynamic range. The ability of GREOL to quantitatively discriminate between isomeric hybrid N-glycans and complex N-glycans in glycoproteins was validated due to the distinct substrate specificities of endoglycosidases. GREOL was also used to analyze changes in human serum N-glycans associated with hepatocellular carcinoma.
稳定同位素标记结合质谱分析是检测聚糖定量变化的一种很有前途的方法，这种方法可能导致许多疾病和紊乱中出现异常糖基化。尽管已经使用了各种同位素标记方法来进行相对糖定量，但尚未开发出类似于蛋白酶催化（18）O 标记用于蛋白质组学的酶（18）O 标记，因为这种方法为糖组学提供了类似的优势。在这项研究中，内切糖苷酶将（18）O 掺入到（18）O 水中释放的糖蛋白 N-聚糖的还原末端，从而使聚糖还原末端（18）O 标记（GREOL）成为相对糖定量的一种潜在策略。由于内切糖苷酶具有不同的底物特异性，该方法在 2 个数量级的动态范围内提供了良好的线性和高重复性。GREOL 还用于分析与肝细胞癌相关的人血清 N-聚糖的变化。
