Glycobiology Advance Access originally published online on March 19, 2003
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Glycobiology, 2003, Vol. 13, No. 8 581-589
© 2003 Oxford University Press
An N-acetylglucosaminyltransferase of the Golgi apparatus of the yeast Saccharomyces cerevisiae that can modify N-linked glycans
MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK
Received on January 22, 2003; revised on February 26, 2003; accepted on February 26, 2003
The yeast Saccharomyces cerevisiae is widely regarded as being only capable of producing N-linked glycans with high-mannose structures. To investigate the glycan structures made in different mutant strains, we made use of a reporter protein consisting of a version of hen egg lysozyme that contains a single site for N-linked glycosylation. Mass spectrometry analysis of the attached glycans revealed that a large proportion contained an unexpected extra mass corresponding to a single N-acetylhexosamine residue. In addition, the glycosylated lysozyme was recognized by an N-acetylglucosamine specific lectin. The genome of S. cerevisiae contains an uncharacterized open reading frame, YOR320c, that is related to a known N-acetylglucosaminyltransferase. Deletion of this ORF resulted in the disappearance of the extra mass on the N-linked glycans and loss of lectin binding. We show that the protein encoded by YOR320c (which we term Gnt1p) is localized to the Golgi apparatus and has GlcNAc-transferase activity in vitro. The physiological role of Gnt1p is unclear because mutants lacking the protein show no obvious growth or cell wall defects. Nonetheless, these results indicate that heterologous glycoproteins expressed in yeast can receive N-glycans with structures other than high mannose. In addition, they indicate that the lumen of the yeast Golgi contains UDP-GlcNAc, which may facilitate reconstitution of higher eukaryotic N-glycan processing.
1 Present address: Research Center for Glycoscience, National Institute of Advanced Industrial Science and Technology, AIST Central 6, Higashi, Tsukuba 305-8566, Japan
2 Present address: Lehrstuhl für Zellbiologie und Pflanzenphysiologie, Universität Regensburg, Universitätsstr. 31, D-93040 Regensburg, Germany
3 To whom correspondence should be addressed; e-mail: sean{at}mrc-lmb.cam.ac.uk
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