Conjugation of carrier molecules to oligosaccharides has been used as a method to enhance the immunogenicity of oligosaccharides in host animals. Since bovine serum albumin and keyhole-limpet hemocyanin, which are commonly used for peptide antigens, are relatively ineffective for enhancing the immunogenicity of conjugated oligosaccharides, improved methods using streptavidin, non-toxic mutants of the diphtheria toxin and phosphatidylethanolamine have been investigated. Phosphatidylethanolamine can easily be conjugated with the reducing end of oligosaccharides, which is an advantage over other carrier molecules, but the ability to enhance the immunogenicity of the conjugated oligosaccharide has not been fully verified. We found that conjugates of target oligosaccharides and a ceramide analogue, characterized by having a very long-chain fatty acid, could strongly induce the production of antibodies that respond to the target oligosaccharides on glycoproteins and glycolipids in host mice. By using mice immunized with this conjugate, we successfully obtained a monoclonal antibody that recognizes a sialylated oligosaccharide structure found on the major serum glycoproteins, and confirmed the high affinity and strict specificity against its oligosaccharide epitope on the glycoproteins.

β-(1,3)-Glucan has been used as an anti-cancer drug owing to its immunostimulatory effects. However, the mechanism of the biological activity has yet to be elucidated. Moreover, β-(1,3)-glucans used for the mechanistic studies are heterogeneous mixtures extracted from natural sources. Therefore, structure-defined β-(1,3)-glucans are required to simplify the effect caused by β-(1,3)-glucans in the study of the biological activity, and this requirement prompts the study of chemical synthesis of β-(1,3)-glucans. The glycosylation for formation of the β-(1,3)-linkage is difficult because the reactivity of the 3-hydroxy group is suppressed due to considerable steric hindrance from the protecting group at the 4-position in glucose. Moreover, introduction of protecting groups that can be deprotected selectively on the 3-position as well as the 6-position is required to synthesize β-(1,3)-glucans with a branch at the 6-position, and the synthetic method should be optimized by considering the influence of the combined protective groups on the glycosylation. Recently, β-(1,3)-glucan oligosaccharide synthesis using a 4,6-O-benzylidenated glycosyl donor and acceptor has been studied strenuously, and the reactivity of the acceptor and the stereoselectivity of β-(1,3)-glycosylation were substantially improved. Consequently, linear β-(1,3)-glucan of up to 16 saccharides could be synthesized. There have been reports of a few promising examples of multiple introductions of branch residues to the linear backbone of β-(1,3)-glucan. Moreover, some applied studies using synthetic β-(1,3)-glucan oligosaccharides have already been conducted.

Notch is a transmembrane protein receptor that mediates direct cell–cell interactions and controls various cell-fate specifications in metazoans. The extracellular domain of Notch contains 36 tandem epidermal growth factor (EGF)-like repeats, most of which have O-linked glycan modifications: O-glucose, O-fucose, and O-GlcNAc. The function of these individual glycans in Notch signaling activation has been investigated by elimination of single modification sites and by knockout of individual glycosyltransferases. Single site mutants show weaker phenotypes compared with glycosyltransferases knockouts in Notch signaling activation. Thus, the collaboration between two or more glycan modifications appears to be essential for full Notch activation. In this review, we describe the history of Notch’s glycan modifications, the individual functions of the modifications, and how glycan modifications might collaborate in regulating Notch function.

Conjugation of carrier molecules to oligosaccharides has been used as a method to enhance the immunogenicity of oligosaccharides in host animals. Since bovine serum albumin and keyhole-limpet hemocyanin, which are commonly used for peptide antigens, are relatively ineffective for enhancing the immunogenicity of conjugated oligosaccharides, improved methods using streptavidin, non-toxic mutants of the diphtheria toxin and phosphatidylethanolamine have been investigated. Phosphatidylethanolamine can easily be conjugated with the reducing end of oligosaccharides, which is an advantage over other carrier molecules, but the ability to enhance the immunogenicity of the conjugated oligosaccharide has not been fully verified. We found that conjugates of target oligosaccharides and a ceramide analogue, characterized by having a very long-chain fatty acid, could strongly induce the production of antibodies that respond to the target oligosaccharides on glycoproteins and glycolipids in host mice. By using mice immunized with this conjugate, we successfully obtained a monoclonal antibody that recognizes a sialylated oligosaccharide structure found on the major serum glycoproteins, and confirmed the high affinity and strict specificity against its oligosaccharide epitope on the glycoproteins.

β-(1,3)-Glucan has been used as an anti-cancer drug owing to its immunostimulatory effects. However, the mechanism of the biological activity has yet to be elucidated. Moreover, β-(1,3)-glucans used for the mechanistic studies are heterogeneous mixtures extracted from natural sources. Therefore, structure-defined β-(1,3)-glucans are required to simplify the effect caused by β-(1,3)-glucans in the study of the biological activity, and this requirement prompts the study of chemical synthesis of β-(1,3)-glucans. The glycosylation for formation of the β-(1,3)-linkage is difficult because the reactivity of the 3-hydroxy group is suppressed due to considerable steric hindrance from the protecting group at the 4-position in glucose. Moreover, introduction of protecting groups that can be deprotected selectively on the 3-position as well as the 6-position is required to synthesize β-(1,3)-glucans with a branch at the 6-position, and the synthetic method should be optimized by considering the influence of the combined protective groups on the glycosylation. Recently, β-(1,3)-glucan oligosaccharide synthesis using a 4,6-O-benzylidenated glycosyl donor and acceptor has been studied strenuously, and the reactivity of the acceptor and the stereoselectivity of β-(1,3)-glycosylation were substantially improved. Consequently, linear β-(1,3)-glucan of up to 16 saccharides could be synthesized. There have been reports of a few promising examples of multiple introductions of branch residues to the linear backbone of β-(1,3)-glucan. Moreover, some applied studies using synthetic β-(1,3)-glucan oligosaccharides have already been conducted.

Notch is a transmembrane protein receptor that mediates direct cell–cell interactions and controls various cell-fate specifications in metazoans. The extracellular domain of Notch contains 36 tandem epidermal growth factor (EGF)-like repeats, most of which have O-linked glycan modifications: O-glucose, O-fucose, and O-GlcNAc. The function of these individual glycans in Notch signaling activation has been investigated by elimination of single modification sites and by knockout of individual glycosyltransferases. Single site mutants show weaker phenotypes compared with glycosyltransferases knockouts in Notch signaling activation. Thus, the collaboration between two or more glycan modifications appears to be essential for full Notch activation. In this review, we describe the history of Notch’s glycan modifications, the individual functions of the modifications, and how glycan modifications might collaborate in regulating Notch function.