title: WbgL: a novel bacterial α1,2-fucosyltransferase for the synthesis of 2'-fucosyllactose.
abstract: Fucosyltransferases (FucTs) are essential tools for the synthesis of fucosylated glycoconjugates. Multistep enzyme catalysis of fucosylated glycans is not limited as long as isolated and well-characterized FucTs are available. The present paper introduces a novel bacterial α1,2-FucT of the glycosyltransferase family 11 encoded by the gene wbgL in the E. coli O126 genome, which only displays 25-30% homology to previously published α1,2-FucTs. A tailor made cloning and expression strategy allowed the successful production of active soluble enzyme in the cytoplasm of E. coli BL21(DE3) and E. coli JM109(DE3), respectively. The lack of a DxD motif and its high activity without divalent metal ions suggests that WbgL belongs to the GT-B fold superfamily. Substrate screening revealed the highest activity for β4-linked galactoside acceptor substrates, such as lactose and lactulose, making WbgL unique among other characterized α1,2-FucTs. Based on its excellent kinetic efficiency for lactose, we present here a sequential reaction strategy for the synthesis of α1,2-fucosyllactose in one pot including the synthesis of the donor substrate 3,3'-Diaminobenzidine (GDP)-β-l-fucose by the bifunctional l-fucokinase/GDP-β-l-Fuc pyrophosphorylase of Bacteroides fragilis 9343.
authors.affiliation: Laboratory for Biomaterials, Institute of Biotechnology and Helmholtz-Institute for Biomedical Engineering, RWTH Aachen University, Worringer Weg 1, 52074 Aachen, Germany.
authors.forename: Leonie
authors.initials: L
authors.lastname: Engels
authors.forename: Lothar
authors.initials: L
authors.lastname: Elling
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001441:Bacteroides fragilis / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005647:Fucosyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006154:Guanosine Diphosphate Fucose / Q000378:metabolism; D007785:Lactose / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D014312:Trisaccharides / Q000096:biosynthesis*
chemical_list: D001426:Bacterial Proteins; D014312:Trisaccharides; D006154:Guanosine Diphosphate Fucose; D005647:Fucosyltransferases; D007785:Lactose; C031420:2'-fucosyllactose
keywords: biocatalysis; glycan epitope; glycoconjugates; glycosyltransferases; nucleotide sugar