Differential Quantitative Determination of Site-Specific Intact N-Glycopeptides in Serum Haptoglobin between Hepatocellular Carcinoma and Cirrhosis Using LC-EThcD-MS/MS
| Autor: | Jie Zhang, David M. Lubman, Qing Yu, St John Skilton, Jianhui Zhu, Lingjun Li, Marshall Bern, Mingrui An, K. Ilker Sen, Zhengwei Chen, Jing Wu |
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| Rok vydání: | 2018 |
| Předmět: |
Liver Cirrhosis
Proteomics 0301 basic medicine medicine.medical_specialty Carcinoma Hepatocellular Glycosylation Cirrhosis Biochemistry Gastroenterology Article Workflow 03 medical and health sciences Tandem Mass Spectrometry Internal medicine medicine Early Hepatocellular Carcinoma Serum haptoglobin Binding Sites Haptoglobins biology business.industry Liver Neoplasms Haptoglobin Glycopeptides General Chemistry medicine.disease Trypsin Glycopeptide Quantitative determination 030104 developmental biology Hepatocellular carcinoma biology.protein business Chromatography Liquid medicine.drug |
| Zdroj: | Journal of Proteome Research. |
| ISSN: | 1535-3907 1535-3893 |
| DOI: | 10.1021/acs.jproteome.8b00654 |
| Popis: | Intact N-glycopeptide analysis remains challenging due to the complexity of glycopeptide structures, low abundance of glycopeptides in protein digests, and difficulties in data interpretation/quantitation. Herein, we developed a workflow that involved advanced methodologies, the EThcD-MS/MS fragmentation method and data interpretation software, for differential analysis of the microheterogeneity of site-specific intact N-glycopeptides of serum haptoglobin between early hepatocellular carcinoma (HCC) and liver cirrhosis. Haptoglobin was immunopurified from 20 μL of serum in patients with early HCC, liver cirrhosis, and healthy controls, respectively, followed by trypsin/GluC digestion, glycopeptide enrichment, and LC-EThcD-MS/MS analysis. Identification and differential quantitation of site-specific N-glycopeptides were performed using a combination of Byonic and Byologic software. In total, 93, 87, and 68 site-specific N-glycopeptides were identified in early HCC, liver cirrhosis, and healthy controls, respectively, with high confidence. The increased variety of N-glycopeptides in liver diseases compared to healthy controls was due to increased branching with hyper-fucosylation and sialylation. Differential quantitation analysis showed that 5 site-specific N-glycopeptides on sites N184 and N241 were significantly elevated in early HCC compared to cirrhosis ( p < 0.05) and normal controls ( p ≤ 0.001). The result demonstrates that the workflow provides a strategy for detailed profiles of N-glycopeptides of patient samples as well as for relative quantitation to determine the level changes in site-specific N-glycopeptides between disease states. |
| Databáze: | OpenAIRE |
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