title: Identification and functional characterization of a highly divergent N-acetylglucosaminyltransferase I (TbGnTI) in Trypanosoma brucei.
abstract: Trypanosoma brucei expresses a diverse repertoire of N-glycans, ranging from oligomannose and paucimannose structures to exceptionally large complex N-glycans. Despite the presence of the latter, no obvious homologues of known β1-4-galactosyltransferase or β1-2- or β1-6-N-acetylglucosaminyltransferase genes have been found in the parasite genome. However, we previously reported a family of putative UDP-sugar-dependent glycosyltransferases with similarity to the mammalian β1-3-glycosyltransferase family. Here we characterize one of these genes, TbGT11, and show that it encodes a Golgi apparatus resident UDP-GlcNAc:α3-D-mannoside β1-2-N-acetylglucosaminyltransferase I activity (TbGnTI). The bloodstream-form TbGT11 null mutant exhibited significantly modified protein N-glycans but normal growth in vitro and infectivity to rodents. In contrast to multicellular organisms, where the GnTI reaction is essential for biosynthesis of both complex and hybrid N-glycans, T. brucei TbGT11 null mutants expressed atypical "pseudohybrid" glycans, indicating that TbGnTII activity is not dependent on prior TbGnTI action. Using a functional in vitro assay, we showed that TbGnTI transfers UDP-GlcNAc to biantennary Man3GlcNAc2, but not to triantennary Man5GlcNAc2, which is the preferred substrate for metazoan GnTIs. Sequence alignment reveals that the T. brucei enzyme is far removed from the metazoan GnTI family and suggests that the parasite has adapted the β3-glycosyltransferase family to catalyze β1-2 linkages.
authors.affiliation: From the Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom and.
authors.forename: Manuela
authors.initials: M
authors.lastname: Damerow
authors.forename: Joao A
authors.initials: JA
authors.lastname: Rodrigues
authors.forename: Di
authors.initials: D
authors.lastname: Wu
authors.forename: M Lucia S
authors.initials: ML
authors.lastname: Güther
authors.forename: Angela
authors.initials: A
authors.lastname: Mehlert
authors.forename: Michael A J
authors.initials: MA
authors.lastname: Ferguson
mesh_terms: D001769:Blood / Q000469:parasitology; D002460:Cell Line; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D009154:Mutation; D017351:N-Acetylglucosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D011134:Polysaccharides / Q000096:biosynthesis; D021381:Protein Transport; D013379:Substrate Specificity; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000502:physiology; D014530:Uridine Diphosphate / Q000378:metabolism
chemical_list: D011134:Polysaccharides; D014530:Uridine Diphosphate; D017351:N-Acetylglucosaminyltransferases; C024850:alpha-1,3-mannosyl-glycoprotein beta-1,2-N-acetylglucosaminyltransferase I
keywords: Glycobiology; Glycosyltransferases; N-Acetylglucosamine; Parasite; Post-translational Modification; Trypanosoma brucei