Transgenic Insect Cell Lines That Support Production of Humanized Glycoproteins By Baculovirus Expression Vectors
Posted on: Thursday, 10 June 2004, 06:00 CDT
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One widely-recognized advantage of the baculovirus-insect cell system is its ability to produce recombinant proteins with cucaryotic modifications, such as glycosylation. However, the protein glycosylation pathways of insect cells appear to be less extensive than those of mammalian cells and, as a result, insect cells rarely produce sialylated glycoproteins. The inability of the baculovirus-inscct cell system to produce authentic recombinant glycoproteins, with carbohydrate side chains identical to those found on native human glycoproteins, can be a serious problem because these side chains can influence glycoprotein behavior in various ways. For the past several years, our group has been creating transgenic lepidopteran insect cell lines with extended protein glycosylation pathways to address this problem. Sf9 and Tn- 5Bl-4 cells were transformed with expression plasmids that encode various mammalian enzymes involved in protein glycosylation. Expression of these genes was controlled using baculovirus enhancer and promoter elements because they provide constitutive foreign gene expression in the absence of baculoviral infection. Clones containing stably integrated copies of these genes were isolated and extensively characterized. Generally, the resulting insect cell lines constitutively expressed the enzymatic activities encoded by the mammalian transgenes. These cells typically had normal morphologies, growth properties, and could support baculovirus infection and recombinant protein production. Furthermore, these transgenic insect cell lines could routinely produce sialylated recombinant glycoproteins under appropriate conditions. Additional details regarding the isolation and characterization of these cell lines will be discussed in this presentation, which will show that we have created transgenic insect cell lines that can support humanized recombinant glycoprotein production by baculovirus expression vectors.
D. L. JARVIS, J. R. Hollister, and J. J. Aumiller. Department of Molecular Biology, University of Wyoming, Laramie, WY 82071. E- mail: dljarvis@uwyo.edu
Copyright Society for In Vitro Biology Spring 2004
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