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Glycobiology Advance Access published online on February 20, 2003
Glycobiology, doi:10.1093/glycob/cwg051
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Submitted on December 26, 2002
Accepted on January 24, 2003

© 2003 Oxford University Press

ORIGINAL ARTICLES

A transgenic insect cell line engineered to produce CMP-sialic acid and sialylated glycoproteins

Jared J. Aumiller 1, Jason R. Hollister 1, Donald L. Jarvis 1*

1 Department of Molecular Biology, University of Wyoming, P.O. Box 3944, Laramie, Wyoming 82071-3944

* To whom correspondence should be addressed. E-mail: DLJarvis{at}uwyo.edu.

Abstract

We previously engineered transgenic insect cell lines to express mammalian glycosyltransferases and showed that these cells can sialylate N-glycoproteins, despite the fact that they have little intracellular sialic acid and no detectable CMP-sialic acid. In the accompanying study, we presented evidence that these cell lines can salvage sialic acids for de novo glycoprotein sialylation from extracellular sialoglycoproteins, such as fetuin, found in fetal bovine serum. This finding led us to create a new transgenic insect cell line designed to synthesize its own sialic acid and CMP-sialic acid. SfSWT-1 cells, which encode five mammalian glycosyltransferases, were transformed with two additional mammalian genes, which encode sialic acid synthase and CMP-sialic acid synthetase. The resulting cell line expressed all seven mammalian genes, produced CMP-sialic acid, and sialylated a recombinant glycoprotein when cultured in a serum-free growth medium supplemented with N-acetylmannosamine. Thus, the addition of mammalian genes encoding two enzymes involved in CMP-sialic acid biosynthesis yielded a new transgenic insect cell line, SfSWT-3, that can sialylate recombinant glycoproteins in the absence of fetal bovine serum. This new cell line will be widely useful as an improved host for baculovirus-mediated recombinant glycoprotein production.


protein glycosylation, insect cells, baculovirus expression system, sialic acid synthesis, genetic engineering, cell transformation
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