A Lactose-Selective α1,3-Fucosyltransferase from Akkermansia muciniphila Expands the Biosynthetic Toolkit for Fucosylated Human Milk Oligosaccharides

Abstract

Fucosylated human milk oligosaccharides (HMOs) are important bioactive glycans, but their biosynthetic production requires fucosyltransferases with predictable acceptor selectivity to avoid regioisomer formation and downstream purification burdens. Here we identify an α1,3-fucosyltransferase from Akkermansia muciniphila (AkkFT) that functions as a lactose-selective α1,3-fucosylation module for HMO synthesis. AkkFT preferentially acts on glycans containing a lactose unit and shows no detectable conversion of type-1 or type-2 N-acetyllactosamine acceptors under standard conditions, contrasting with commonly used bacterial α1,3/4-fucosyltransferases. This lactose-anchored selectivity enables late-stage α1,3-fucosylation across neutral and sialylated HMO acceptors, including efficient formation of lacto-N-neofucopentaose V and the blood group B antigen. Notably, AkkFT converts disialyl-lacto-N-tetraose to disialyl-lacto-N-fucopentaose V under conditions where Helicobacter pylori α1,3-fucosyltransferases show no detectable activity. AkkFT also converts free 2′-fucosyllactose to lactodifucotetraose on a preparative scale and was implemented in an engineered Escherichia coli strain for whole-cell 3-fucosyllactose production. Overall, AkkFT expands the repertoire of selectivity-defined biosynthetic parts for fucosylated HMO production and demonstrates how enzyme discovery can complement engineered microbial platforms for glycan biomanufacturing.