A chemical synthesis of a core fucose containing N-glycan was achieved. Asparagine was introduced at an early stage of the synthesis, and the sugar chain was convergently elongated. As for the fragment synthesis, we reinvestigated α-sialylation, β-mannosylation, and N-glycosylation to reveal that precise temperature control was essential for these glycosylations. Intermolecular hydrogen bonds involving acetamide groups were found to reduce the reactivity in glycosylations: the protection of NHAc as NAc dramatically improved the reactivity. The dodecasaccharide-asparagine framework was constructed via the (4 + 4) glycosylation and the (4 + 8) glycosylation using the tetrasaccharide donor and the tetrasaccharide-asparagine acceptor. An ether-type solvent enhanced the yields of these key glycosylations between large substrates. After the whole deprotection of the dodecasaccharide, the target N-glycan was obtained.

The chemical synthesis of a bisecting N-acetylglucosamine (GlcNAc)-containing N-glycan was achieved by a convergent synthetic route through [4+2] and [6+2] glycosylations. This synthetic route reduced the number of reaction steps, although the key glycosylations were challenging in terms of yields and selectivities owing to steric hindrance at the glycosylation site and a lack of neighboring group participation. The yields of these glycosylations were enhanced by stabilizing the oxocarbenium ion intermediate through ether coordination. Glycosyl donor protecting groups were explored in an effort to realize perfect α selectivity by manipulating remote participation. The simultaneous glycosylations of a tetrasaccharide with two disaccharides was investigated to efficiently construct a bisecting GlcNAc-containing N-glycan.

We have elucidated the cooperative stabilization of proteins by sugars, amino acids, and other protein-stabilizing agents using a new and simple assay system. Our system determines the protein-stabilizing ability of various compounds by measuring their ability to protect hemoglobin from denaturation. Hemoglobin denaturation was readily measured by quantitative changes in its ultraviolet-visible absorption spectrum. The efficiency of our assay was confirmed using various sugars such as trehalose and sucrose that are known to be good protein stabilizers. We have also found that mixtures of two different types of protein stabilizers resulted in a cooperative stabilizing effect on protein.