title: Identification of sialylated glycoproteins from metabolically oligosaccharide engineered pancreatic cells.
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.
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Tian
authors.affiliation: Department of Biomedical Engineering and the Translational Tissue Engineering Center, Johns Hopkins University School of Medicine, Baltimore, MD USA.
authors.forename: Ruben T
authors.initials: RT
authors.lastname: Almaraz
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Caitlin H
authors.initials: CH
authors.lastname: Choi
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Qing Kay
authors.initials: QK
authors.lastname: Li
authors.affiliation: Department of Biomedical Engineering and the Translational Tissue Engineering Center, Johns Hopkins University School of Medicine, Baltimore, MD USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Saeui
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Danni
authors.initials: D
authors.lastname: Li
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Punit
authors.initials: P
authors.lastname: Shah
authors.affiliation: Department of Biomedical Engineering and the Translational Tissue Engineering Center, Johns Hopkins University School of Medicine, Baltimore, MD USA.
authors.forename: Rahul
authors.initials: R
authors.lastname: Bhattacharya
authors.affiliation: Department of Biomedical Engineering and the Translational Tissue Engineering Center, Johns Hopkins University School of Medicine, Baltimore, MD USA.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Yarema
authors.affiliation: Department of Pathology, Johns Hopkins University, 400 N. Broadway, Room 4011, Baltimore, MD 21287 USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
keywords: Metabolic oligosaccharide engineering; Pancreatic cancer cells; Sialylated glycoproteins