원문정보
초록
영어
The characterization of site-specific micro-heterogeneity in glycoprotein is very important for understanding cell biology and disease processes. Vitronectin is well known to be a multi-functional glycoprotein in blood and the extracellular matrix, which is related to hepatocellular carcinoma (HCC). Here, we systematically analyzed the site-specific N-glycopeptides of vitronectin in human plasma by tandem mass spectrometry combined with immunoprecipitation and HILIC enrichment. Vitronectin was purified with immunoprecipitation by monoclonal antibody from plasma and digested to tryptic N-glycopeptides. Then, enrichment with HILIC materials was used, and followed by analysis with nano-LC/MS/MS. The sequences of N-glycopeptides were identified from the mass spectra by high-energy C-trap dissociation (HCD) and collision-induced dissociation (CID). In HCD mode, oxonium ions were used for recognizing glycopeptides and y ions for sequencing the peptide backbone. In CID mode, Y ions were used for characterizing their glycoforms. As a result, total 17 site-specific N-glycopeptides were completely identified at all of three N-glycosylation sites of vitronectin in human plasma, including 12 N-glycopeptides first reported. Finally, we specifically found that three hybrid and four complex glycopeptides of tri-antennary forms with outer-fucosylation increased in HCC human plasma.