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US 10,538,794 B2Granted
Oligosaccharide libraries and methods of production
AssigneeEmory University
InventorsTongzhong Ju, Richard Cummings, Matthew Kudelka
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Abstract
This disclosure relates to libraries of oligosaccharides comprising multiple individual oligosaccharides derived from cells that contain a tag or detectable marker. In certain embodiments, the tag or detectable marker is an oxygen linked to the oligosaccharides through the first carbon of a sugar ring. In certain embodiments, the disclosure relates to methods of generating the libraries of oligosaccharides using saccharides, e.g., monosaccharides, or a disaccharide, in which hydroxyl groups are chemically acetylated.
Claims7
A method of creating and analyzing a library of oligosaccharides comprising,
- (i) providing an acetylated tagged oxygen linked saccharide, wherein the acetylated tagged oxygen linked saccharide is benzyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-α-D-galactopyranoside;
- (ii) mixing the acetylated tagged oxygen linked saccharide with a living in vitro cell at a concentration of less than 250 micromolar (μM) under conditions such that the acetylated tagged oxygen linked saccharide penetrates into the living in vitro cell, and under conditions such that the acetylated tagged oxygen linked saccharide is converted into multiple tagged oxygen linked oligosaccharides within an incubation time of three (3) days;
- (iii) obtaining the multiple tagged oxygen linked oligosaccharides that are secreted from the living in vitro cell or that are present in a cellular cytosol of the living in vitro cell;
- (iv) separating the multiple tagged oxygen linked oligosaccharides to provide the library of oligosaccharides comprising purified individual tagged oxygen linked oligosaccharides;
- and
- (v) analyzing the purified individual tagged oxygen linked oligosaccharides by mass spectroscopy to provide measured molecular weights of the purified individual tagged oxygen linked oligosaccharides to provide an O-glycome profile.
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Dependencies are inferred from the claim text.