Peptidoglycan (PGN) is a component of bacterial cell walls; its fragments are recognized by the cytoplasmic receptors Nod1 and Nod2, thereby promoting the production of inflammatory cytokines and antibodies. To further elucidate these biological defense mechanisms, a large and stable supply of the PGN fragments via chemical synthesis is essential. However, the synthesis and purification of long PGN fragments are quite challenging due to their low solubility. In this study, we efficiently synthesized PGN fragments via solid-phase oligosaccharide synthesis (SPOS). Using the JandaJel™ Wang resin (JJ-Wang), an octasaccharide glycan chain of PGN was constructed by repeating glycosylation reactions to elongate β-1,4-linked disaccharide units composed of MurNAc and GlcNAc. To enhance reactivity, glycosylation was performed in a mixed solvent comprising C4F9OEt/CH2Cl2/THF with the intention of promoting substrate concentration onto the solid support through the fluorophobic effect, affording the PGN octasaccharide in a 19% overall yield (10 steps). Subsequently, after deprotection of the O-Fmoc, N-Troc, and ethyl ester groups, N- and O-acetylation proceeded smoothly, owing to the high swelling property of JJ-Wang. Peptide condensation with L-Ala-D-isoGln(OBn) and carboxylic acids was also achieved. Finally, cleavage of the PGN fragment from the resin with TFA afforded the desired octasaccharide with dipeptides in a 2.3% overall yield (15 steps).
Keywords: glycosylation; innate immunity; peptidoglycan; solid-phase oligosaccharide synthesis.