title: Improved glycosylation of a foreign protein by Tn-5B1-4 cells engineered to express mammalian glycosyltransferases.
abstract: The major advantages of using the baculovirus-insect cell system for recombinant protein production are its ability to produce large amounts of recombinant proteins and its ability to provide eucaryotic modifications, such as glycosylation. However, the glycans linked to recombinant glycoproteins produced by this system typically differ from those found on native mammalian products. This is an important problem because glycans on mammalian glycoproteins can influence their functions in many different ways. The inability of baculovirus-infected insect cells to produce glycans identical to those found on native mammalian glycoproteins is due, in part, to the absence of functional levels of certain glycosyltransferases in insect cells. Thus, the purpose of this study was to engineer these activities into Tn-5B1-4, an established insect cell line that is widely used as a host for baculovirus-mediated protein production. Expression plasmids were constructed in which cDNAs encoding mammalian beta1,4-galactosyltransferase and alpha2,6-sialyltransferase were placed under the transcriptional control of a baculovirus immediate early promoter. These plasmids were then used to isolate two different transgenic Tn-5B1-4 derivatives and the biological and biochemical properties of these cell lines were examined. The results show that both of the engineered insect cell lines have improved glycoprotein-processing capabilities, relative to the parental cell line.
authors.affiliation: Department of Molecular Biology, University of Wyoming, P.O. Box 3944, Laramie, WY 82071-3944, USA.
authors.forename: K
authors.initials: K
authors.lastname: Breitbach
authors.forename: D L
authors.initials: DL
authors.lastname: Jarvis
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016367:Baculoviridae / Q000235:genetics; D002478:Cells, Cultured; D006031:Glycosylation; D016695:Glycosyltransferases / Q000235:genetics* / Q000378:metabolism; D008322:Mammals / Q000235:genetics*; D009036:Moths / Q000166:cytology*; D009237:N-Acetyllactosamine Synthase / Q000235:genetics / Q000378:metabolism; D009355:Neomycin / Q000494:pharmacology; D010957:Plasmids / Q000187:drug effects; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012799:Sialyltransferases / Q000235:genetics / Q000378:metabolism; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
chemical_list: D000900:Anti-Bacterial Agents; D011994:Recombinant Proteins; D016695:Glycosyltransferases; D009237:N-Acetyllactosamine Synthase; D012799:Sialyltransferases; D009355:Neomycin; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase