Shotgun Glycomics: A Microarray Strategy for Functional Glycomics

• Authors: Xuezheng Song, Emory University; Yi Lasanajak, Emory University; Baoyun Xia, Emory University; Jamie Heimburg-Molinaro, Emory University; Jeanne M. Rhea, Emory University; Hong Ju, Emory University; Chunmei Zhao, Emory University; Ross J Molinaro, Emory University; Richard D. Cummings, Emory University; David Smith, Emory University
• Journal Title: Nature Methods
• Volume: Volume 8, Number 1
• Publisher: Nature Research (part of Springer Nature) | 2011-01, Pages 85-90
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1038/nmeth.1540
• Subjects: Chemistry, Biochemistry
• Keywords: glycosphingolipids; glycan array; fluorescent labeling; immobilization; functional glycomics

Abstract

Major challenges of glycomics are to characterize a glycome and identify functional glycans as ligands for glycan-binding proteins (GBPs). To address these issues we have developed a general strategy termed shotgun glycomics. We focus on glycosphingolipids (GSLs), a challenging class of glycoconjugates recognized by toxins, antibodies, and GBPs. We derivatized GSLs extracted from cells with a heterobifunctional fluorescent tag suitable for covalent immobilization. Fluorescent GSLs were separated by multidimensional chromatography, quantified, and coupled to glass slides to create GSL shotgun microarrays. The microarrays were interrogated with cholera toxin, antibodies, and sera from patients with Lyme disease to identify biologically relevant GSLs that were subsequently characterized by mass spectrometry. Shotgun glycomics incorporating GSLs and potentially glycoprotein-derived glycans provides an approach to accessing the complex glycomes of animal cells and offers a strategy for focusing structural analyses on functionally significant glycans.