PNGase F-mediated incorporation of (18)O into glycans for relative glycan quantitation

• Published in: Analyst (London) Vol. 140; no. 4; pp. 1082 - 1089
• Main Authors: Zhang, Wei, Cao, Weiqian, Huang, Jiangming, Wang, Hong, Wang, Ji, Xie, Chen, Yang, Pengyuan
• Format: Journal Article
• Language: English
• Published: England 21.02.2015
• Subjects: Amino Acid Sequence; Carbohydrate Sequence; Carcinoma, Hepatocellular - metabolism; Glycomics - methods; Glycopeptides - chemistry; Glycopeptides - metabolism; Glycoproteins - chemistry; Glycoproteins - metabolism; Glycosylation; Humans; Immunoglobulin G - chemistry; Immunoglobulin G - metabolism; Liver Neoplasms - metabolism; Mass Spectrometry - methods; Molecular Sequence Data; Oxygen Isotopes - analysis; Oxygen Isotopes - metabolism; Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase - metabolism; Polysaccharides - analysis; Polysaccharides - metabolism; Reproducibility of Results
• ISSN: 1364-5528
• 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.