Glycobiology Advance Access published online on December 22, 2004
Glycobiology, doi:10.1093/glycob/cwi032
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
1 Mammalian Cell Technology, Novo Nordisk A/S, Novo Allé, 2880 Bagsværd, Denmark
* To whom correspondence should be addressed. N-glycosylation is normally a co-translational process, which occurs during translocation of the nascent protein to the endoplasmic reticulum. In the present study, however, we demon-strate post-translational N-glycosylation of recombinant human coagulation factor VII (FVII) in CHO-K1 and 293A cells. Human FVII has two N-glycosylation sites (N145 and N322). Pulse-chase labelled intracellular FVII migrated as two bands corresponding to FVII with one and two N-glycans, respectively. N-glycosidase treatment converted both these band into a single band, which comigrated with mutated FVII without N-glycans. Immediately after pulse, most labelled intracellular FVII had one N-glycan, but during a 1 h chase, the vast ma-jority was processed into FVII with two N-glycans demonstrating post-translational N-glycosylation of FVII. Pulse-chase analysis of N-glycosylation site knock-out mutants de-monstrated co-translational glycosylation of N145 but primarily or exclusively post-translational glycosylation of N322. The post-translational N-glycosylation appeared to take place in the same time-frame as the folding of nascent FVII into a secretion-competent con-formation, indicating a link between the two processes. We propose that the co-translational conformation(s) of FVII are unfavourable for glycosylation at N332, whereas a more favour-able conformation is obtained during the post-translational folding. This is the first documen-tation of post-translational N-glycosylation of a non-modified protein in mammalian cells with an intact N-glycosylation machinery. Thus, the present study demonstrates that post-translational N-glycosylation can be a part of the normal processing of glycoproteins.
Received October 22, 2004
Revised December 15, 2004
Accepted December 15, 2004
Article
Post-translational N-glycosylation takes place during the normal processing of human coagulation factor VII
Gert Bolt, E-mail: bolt{at}novonordisk.com
Abstract 
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  U. R. Mbonye, C. Yuan, C. E. Harris, R. S. Sidhu, I. Song, T. Arakawa, and W. L. Smith Two Distinct Pathways for Cyclooxygenase-2 Protein Degradation J. Biol. Chem., March 28, 2008; 283(13): 8611 - 8623. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Biscetti Glycosylation of Thrombin Activatable Fibrinolysis Inhibitor: Why Is it So Important? Circ. Res., February 15, 2008; 102(3): 278 - 279. [Full Text] [PDF]
|
|
|
|
|
|
R. K. Ambasta, X. Ai, and C. P. Emerson Jr. Quail Sulf1 Function Requires Asparagine-linked Glycosylation J. Biol. Chem., November 23, 2007; 282(47): 34492 - 34499. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-M. Krause, P. Berger, J. Roig, V. Singh, and W. E. Merz Rapid Maturation of Glycoprotein Hormone Free {alpha}-Subunit (GPH{alpha}) and GPH{alpha}{alpha} Homodimers Mol. Endocrinol., October 1, 2007; 21(10): 2551 - 2564. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Kowarik, S. Numao, M. F. Feldman, B. L. Schulz, N. Callewaert, E. Kiermaier, I. Catrein, and M. Aebi N-Linked Glycosylation of Folded Proteins by the Bacterial Oligosaccharyltransferase. Science, November 17, 2006; 314(5802): 1148 - 1150. [Abstract] [Full Text] [PDF]
|
|