Identification of sialylated glycoproteins from metabolically oligosaccharide engineered pancreatic cells

Yuan Tian, Ruben T. Almaraz, Caitlin H. Choi, Qing Kay Li, Christopher Saeui, Danni Li, Punit Shah, Rahul Bhattacharya, Kevin J. Yarema, Hui Zhang

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

In this study, we investigated the use of metabolic oligosaccharide engineering and bio-orthogonal ligation reactions combined with lectin microarray and mass spectrometry to analyze sialoglycoproteins in the SW1990 human pancreatic cancer line. Specifically, cells were treated with the azido N-acetylmannosamine analog, 1,3,4-Bu3ManNAz, to label sialoglycoproteins with azide-modified sialic acids. The metabolically labeled sialoglyproteins were then biotinylated via the Staudinger ligation, and sialoglycopeptides containing azido-sialic acid glycans were immobilized to a solid support. The peptides linked to metabolically labeled sialylated glycans were then released from sialoglycopeptides and analyzed by mass spectrometry; in parallel, the glycans from azido-sialoglycoproteins were characterized by lectin microarrays. This method identified 75 unique N-glycosite-containing peptides from 55 different metabolically labeled sialoglycoproteins of which 42 were previously linked to cancer in the literature. A comparison of two of these glycoproteins, LAMP1 and ORP150, in histological tumor samples showed overexpression of these proteins in the cancerous tissue demonstrating that our approach constitutes a viable strategy to identify and discover sialoglycoproteins associated with cancer, which can serve as biomarkers for cancer diagnosis or targets for therapy.

Original languageEnglish (US)
Article number11
JournalClinical Proteomics
Volume12
Issue number1
DOIs
StatePublished - 2015

Bibliographical note

Funding Information:
This work was supported by federal funds from the National Cancer Institute, National Institutes of Health, grants 2R01CA112315, U01CA152813, R01CA112314, and P01HL107153-01. We also gratefully acknowledge the support of Drs. Robert Cole and Robert O’Meally from Johns Hopkins University for their assistance in mass spectrometry analysis and data processing.

Publisher Copyright:
© 2015 Tian et al.

Keywords

  • Metabolic oligosaccharide engineering
  • Pancreatic cancer cells
  • Sialylated glycoproteins

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