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Edited by Carolyn R. Bertozzi, University of California, Berkeley, CA, and approved January 7, 2008 (received for review November 1, 2007)
Abstract
Mucin-type O-glycans are the most typical O-glycans found in mammalian cells and assume many different biological roles. Here, we report a genetic engineered yeast strain capable of producing mucin-type sugar chains. Genes encoding Bacillus subtilis UDP-Gal/GalNAc 4-epimerase, human UDP-Gal/GalNAc transporter, human ppGalNAc-T1, and Drosophila melanogaster core1 β1–3 GalT were introduced into Saccharomyces cerevisiae. The engineered yeast was able to produce a MUC1a peptide containing O-glycan and also a mucin-like glycoprotein, human podoplanin (hPod; also known as aggrus), which is a platelet-aggregating factor that requires a sialyl-core1 structure for activity. After in vitro sialylation, hPod from yeast could induce platelet aggregation. Interestingly, substitution of ppGalNAc-T1 for ppGalNAc-T3 caused a loss of platelet aggregation-inducing activity, despite the fact that the sialyl-core1 was detectable in both hPod proteins on a lectin microarray. Most of O-mannosylation, a common modification in yeast, to MUC1a was suppressed by the addition of a rhodanine-3-acetic acid derivative in the culture medium. The yeast system we describe here is able to produce glycoproteins modified at different glycosylation sites and has the potential for use in basic research and pharmaceutical applications.
Footnotes
- †To whom correspondence may be addressed. E-mail: y-chiba{at}aist.go.jp or jigami.yoshi{at}aist.go.jp
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Author contributions: Y.C., J.H., Y.J., and H.N. designed research; K.A., Y. Kasahara, Y. Kato, M.K.K., A.K., H.I., and K.K. performed research; K.A., Y. Kasahara, Y. Kato, and A.K. analyzed data; and K.A., Y.C., and Y. Kato wrote the paper.
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The authors declare no conflict of interest.
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This article is a PNAS Direct Submission.
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This article contains supporting information online at www.pnas.org/cgi/content/full/0710412105/DC1.
- © 2008 by The National Academy of Sciences of the USA
Engineering of mucin-type human glycoproteins in yeast cells
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