Neoglycolipid Technology: Deciphering Information Content of Glycome

Abstract

Publisher Summary Neoglycolipid technology was introduced in 1985 to enable direct binding studies to be conveniently performed with oligosaccharides of glycoproteins. At that time, there was a lack of microscale methods for examining the carbohydrate–protein interactions, after release of the oligosaccharides from the carrier proteins. Most carbohydrate–protein interactions are of such low affinities that di- or multivalence of both oligosaccharide and recognition protein is required for detection by precipitation, radioimmunoassay, or enzyme-linked immunosorbent assay experiments. It was clear that a method was required to examine the released oligosaccharides in a multivalent state. The existing methods for examining the recognition of specific oligosaccharides by antibodies and other carbohydrate-recognizing proteins involved purification of the oligosaccharides, often from highly heterogeneous mixtures, and testing them individually as inhibitors of the binding of the recognition proteins to macromolecules or cells. The amounts of oligosaccharides required for inhibition of binding were frequently prohibitive. The approach chosen to address the problem was chemical conjugation of oligosaccharides to lipid. The artificial glycolipids, neoglycolipids, formed would enable the presentation of the oligosaccharides in the clustered state. The conjugation principle selected was reductive amination to a phosphatidylethanolaminetype aminolipid. For O-glycans released by reductive alkali treatment, a mild periodate oxidation procedure was included to generate reactive aldehydes for the conjugation. A major advantage of conjugating each oligosaccharide in a mixture to a lipid molecule rather than to a protein, such as bovine serum albumin, is that each component remains discrete and can be isolated.