Controllable Enzymatic\nSynthesis of Natural Asymmetric\nHuman Milk Oligosaccharides

Among human milk\noligosaccharides (HMOs), linear HMOs\nare synthesized\nthrough mature but varied routes. Although branched HMOs can be synthesized\nby chemical, enzymatic, or chemoenzymatic methods, these methods cannot\nbe easily applied to the synthesis of asymmetric multiantennary oligosaccharides.\nHerein, we developed a controllable method to synthesize asymmetric\nbiantennary HMOs. In our synthetic route, GlcNAcβ1,3(GlcN3β1,6)Glaβ1,4Glc\nwas first chemically synthesized as the core tetrasaccharide, which\ncontains β1,6GlcN3 as the “stop” sugar in transferase-catalyzed\nglycosylation. The desired sugars at the GlcNAcβ1–3Gal\narm can be assembled using galactosyltransferase, <i>N</i>-acetylglucosaminyltransferase, and fucosyltransferase. Then, the\nStaudinger reduction and acetylation were used to transform GlcN3\nto GlcNAc and assemble sugars by initiating the “go”\nprocess. By manipulating transferase-catalyzed glycosylations, 22\nnatural asymmetric biantennary oligosaccharides were synthesized.