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Glycobiology Advance Access published online on November 1, 2002
Glycobiology, doi:10.1093/glycob/cwg012
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Submitted on July 21, 2002
Revised on August 29, 2002
Accepted on September 5, 2002

© 2002 Oxford University Press

ORIGINAL ARTICLES

Complex-type biantennary N-glycans of recombinant human transferrin from Trichoplusia ni insect cells expressing mammalian {beta}-1,4-galactosyltransferase and {beta}-1,2-N-acetylglucosaminyltransferase II

Noboru Tomiya 1*, Dale Howe 2, Jared J. Aumiller 2, Manuj Pathak 3, Jung Park 4, Karen Palter 4, Donald L. Jarvis 2, Michael J. Betenbaugh 3, Yuan C. Lee 1

1 Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218
2 Department of Molecular Biology, University of Wyoming, Laramie, Wyoming, 82071-3944
3 Department of Chemical Engineering, Johns Hopkins University, Baltimore, Maryland 21218
4 Department of Biology, Temple University, Philadelphia, Pennsylvania, 19122

* To whom correspondence should be addressed. E-mail: ntomiya1{at}jhu.edu.

Abstract

A novel recombinant baculovirus expression vector was used to produce His-tagged human transferrin in a transformed insect cell line (Tn5{beta}4GalT) that constitutively expresses a mammalian {beta}-1,4-galactosyltransferase. This virus encoded the His-tagged human transferrin protein in conventional fashion, under the control of the very late polyhedrin promoter. In addition, to enhance the synthesis of galactosylated biantennary N-glycans, this virus encoded human {beta}-1,2-N-acetylglucosaminyltransferase II under the control of an immediate-early (ie1) promoter. Detailed analyses by MALDI-TOF mass spectrometry, exoglycosidase digestion, and two-dimensional HPLC revealed that the N-glycans on the purified recombinant human transferrin produced by this virus-host system included four different fully galactosylated, biantennary, complex-type glycans. Thus, this study describes a novel baculovirus-host system, which can be used to produce a recombinant glycoprotein with fully galactosylated, bi-antennary N-glycans.


Key words:N-glycan; Insect; HPLC; MALDI-MS
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O. Choi, N. Tomiya, J. H. Kim, J. M. Slavicek, M. J. Betenbaugh, and Y. C. Lee
N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system
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[Abstract] [Full Text] [PDF]


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