Exploring Regioselective Fucosylation Catalyzed by Bacterial Glycosyltransferases through Substrate Promiscuity and Acceptor-Mediated Glycosylation

Fucose in fucosylated glycans isprimarily assembledat multiplesets of N-acetyllactosamine (LN) and lacto-N-biose (LNB) units, which are known as Lewis antigens.The synthesis of these compounds is difficult due to the diversityof Lewis antigens on complex glycans. A promising strategy is to performenzymatic fucosylation with oligosaccharide backbones because themethod is stereoselective and synthetically efficient. However, purificationis difficult with this method because heterogeneous glycans are produced.Herein, we developed a general strategy to perform site-specific fucosylationon poly LN or LN/LNB hybrid backbones. We found that trifluoroacetylglucosamine (GlcNTFA) can function as a GlcNAc substituent and beincorporated into the LN/LNB chain. The TFA group was readily convertedto a butyloxycarbonyl (Boc) group, and the resulting GlcNBoc can beaccepted by bacterial galactosyltransferases and provides steric hindranceto prevent fucosyltransferase recognition. Thus, enzymatic fucosylationcan only occur specifically at GlcNAc sites within GlcNHBoc-containingpoly LN/LNB. We demonstrated the feasibility of this strategy by efficientlysynthesizing myeloglycans (dodecasaccharides) and internal/terminalfucosylated human milk oligosaccharides. This approach offers a solutionto bypass complex chemical glycosylations and control the selectivityof enzymatic fucosylations. Thus, the method shows significant potentialfor the future synthesis of branched glycans, such as N-glycans and O-glycans.