A Bacterial ?1-3-Galactosyltransferase Enables Multigram-Scale Synthesis of Human Milk Lacto-N-tetraose (LNT) and Its Fucosides.

ABSTRACT: ?1-3-Linked galactosides such as Gal?1?3GlcNAc?OR are common carbohydrate motifs found in human milk oligosaccharides (HMOSs), glycolipids, and glycoproteins. Efficient and scalable enzymatic syntheses of these structures have proven challenging due to the lack of access to a highly active ?1?3-galactosyltransferase (?3GalT) in large amounts. Previously reported E. coli ?3GalT (EcWbgO) has been identified as a limiting factor for producing a ?1-3-galactose-terminated human milk oligosaccharide lacto-N-tetraose (LNT) by fermentation. Here we report the identification of an EcWbgO homolog from C. violaceum (Cv?3GalT) which showed a high efficiency in catalyzing the formation of LNT from lacto-N-triose (LNT II). With the highly active Cv?3GalT, multigram-scale (>10 gram) synthesis of LNT from lactose was achieved using a sequential one-pot multienzyme (OPME) glycosylation process. The access to Cv?3GalT enabled enzymatic synthesis of several fucosylated HMOSs with or without further sialylation including LNFP II, S-LNF II, LNDFH I, LNFP V, and DiFuc-LNT. Among these, LNFP V and DiFuc-LNT would not be accessible by enzymatic synthesis if an active ?3GalT were not available.

SUBMITTER: McArthur JB 

PROVIDER: S-EPMC7785058 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature