Array-assisted Characterization of a Fucosyltransferase Required for the Biosynthesis of Complex Core Modifications of Nematode N-Glycans

Fucose is a common monosaccharide component of cell surfaces and is involved in many biologicalrecognition events. Therefore, definition and exploitation of the specificity of the enzymes(fucosyltransferases) involved in fucosylation is a recurrent theme in modern glycosciences. Despitevarious studies, the specificities of many fucosyltransferases are still unknown, so new approachesare required to study these. The model nematode Caenorhabditis elegans expresses awide range of fucosylated glycans, including N-linked oligosaccharides with unusualcomplex core modifications. Up to three fucose residues can be present on the standardN,N′-diacetylchitobiose unit of these N-glycans, but onlythe fucosyltransferases responsible for transfer of two of these (the coreα1,3-fucosyltransferase FUT-1 and the core α1,6-fucosyltransferase FUT-8) werepreviously characterized. By use of a glycan library in both array and solution formats, we wereable to reveal that FUT-6, another C. elegans α1,3-fucosyltransferase,modifies nematode glycan cores, specifically the distal N-acetylglucosamineresidue; this result is in accordance with glycomic analysis of fut-6 mutant worms.This core-modifying activity of FUT-6 in vitro and in vivo is inaddition to its previously determined ability to synthesize Lewis X epitopes invitro. A larger scale synthesis of a nematode N-glycan core invitro using all three fucosyltransferases was performed, and the nature of the glycosidiclinkages was determined by NMR. FUT-6 is probably the first eukaryotic glycosyltransferase whosespecificity has been redefined with the aid of glycan microarrays and so is a paradigm for the studyof other unusual glycosidic linkages in model and parasitic organisms.