Identification of a glycosylphosphatidylinositol anchor-modifying beta1-3 N-acetylglucosaminyl transferase in Trypanosoma brucei.
Trypanosoma brucei expresses complex glycoproteins throughout its life cycle. A review of its repertoire of glycosidic linkages suggests a minimum of 38 glycosyltransferase activities. Of these, five have been experimentally related to specific genes and a further nine can be associated with candidate genes. The remaining linkages have no obvious candidate glycosyltransferase genes; however, the T. brucei genome contains a family of 21 putative UDP sugar-dependent glycosyltransferases of unknown function. One representative, TbGT8, was used to establish a functional characterization workflow. Bloodstream and procyclic-form TbGT8 null mutants were created and both exhibited normal growth. The major surface glycoprotein of the procyclic form, the procyclin, exhibited a marked reduction in molecular weight due to changes in the procyclin glycosylphosphatidylinositol (GPI) anchor side-chains. Structural analysis of the mutant procyclin GPI anchors indicated that TbGT8 encodes a UDP-GlcNAc: beta-Gal-GPI beta1-3 GlcNAc transferase. This is only the second GPI-modifying glycosyltransferase to have been identified from any organism. The glycosylation of the major glycoprotein of bloodstream-form T. brucei, the variant surface glycoprotein, was unaffected in the TbGT8 mutant. However, changes in the lectin binding of other glycoproteins suggest that TbGT8 influences the processing of the poly N-acetyllactosamine-containing asparagine-linked glycans of this life cycle stage.
Izquierdo
Luis
L
Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
Nakanishi
Masayuki
M
Mehlert
Angela
A
Machray
Greg
G
Barton
Geoffrey J
GJ
Ferguson
Michael A J
MA
D000818:Animals; D016054:DNA, Protozoan / Q000235:genetics; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D017351:N-Acetylglucosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D014170:Transformation, Genetic; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000378:metabolism*; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000378:metabolism
D016054:DNA, Protozoan; C406750:EP procyclin protein, Trypanosoma brucei; C406751:GPEET protein, Trypanosoma brucei; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D015800:Protozoan Proteins; D014643:Variant Surface Glycoproteins, Trypanosoma; D017351:N-Acetylglucosaminyltransferases