Cellular O-Glycome Reporter/Amplification to explore O-glycans of living cells

• Authors: Matthew R. Kudelka, Emory University; Aristotelis Antonopoulos, Imperial College London; Yingchun Wang, Emory University; Duc M. Duong, Emory University; Xuezheng Song, Emory University; Nicholas Seyfried, Emory University; Anne Dell, Imperial College London; Stuart M. Haslam, Imperial College London; Richard Cummings, Emory University; Tongzhong Ju, Emory University
• Journal Title: Nature Methods
• Volume: Volume 13, Number 1
• Publisher: Nature Publishing Group | 2016-01-01, Pages 81-+
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1038/NMETH.3675
• Keywords: Science & Technology; Life Sciences & Biomedicine; Biochemical Research Methods; Biochemistry & Molecular Biology; SIALYL-LEWIS-X; PLURIPOTENT STEM-CELLS; MASS-SPECTROMETRY; IN-VITRO; CANCER; GLYCOSYLATION; TISSUES; DISEASE; ANTIGEN; COSMC; Carbohydrates; Glycobiology; Mass spectrometry

Abstract

Protein O-glycosylation has key roles in many biological processes, but the repertoire of O-glycans synthesized by cells is difficult to determine. Here we describe an approach termed Cellular O-Glycome Reporter/Amplification (CORA), a sensitive method used to amplify and profile mucin-type O-glycans synthesized by living cells. Cells convert added peracetylated benzyl-α-N-acetylgalactosamine to a large variety of modified O-glycan derivatives that are secreted from cells, allowing for easy purification for analysis by HPLC and mass spectrometry (MS). Relative to conventional O-glycan analyses, CORA resulted in an ∼100-1,000-fold increase in sensitivity and identified a more complex repertoire of O-glycans in more than a dozen cell types from Homo sapiens and Mus musculus. Furthermore, when coupled with computational modeling, CORA can be used for predictions about the diversity of the human O-glycome and offers new opportunities to identify novel glycan biomarkers for human diseases.