ISSN:
1573-4986
Keywords:
O-glycosylation signal
;
mucin
;
fibroblast growth factor
;
signal sequence
;
CHO cells, Chinese hamster ovary cells
;
FGF, fibroblast growth factor
;
Gal, galactose
;
GalNAc, N-acetylgalactosamine
;
PNGase, peptide N-glycosidase F
;
HUVEC, human umbilical vein endothelial cell
Source:
Springer Online Journal Archives 1860-2000
Topics:
Chemistry and Pharmacology
Notes:
Abstract The peptide signal sequence for protein O-glycosylation is not fully characterized, although a recent in vitro study proposed that the sequence motif, XTPXP, serves as a signal for mucin-type O-glycosylation. Here, we show that the AATPAP sequence acts as an efficient O-glycosylation signal, in vivo. A secreted fibroblast growth factor (secFGF) was used as a model to analyze glycosylation and its effects on the biological activity of FGF. Two constructs encoding [AATPAP]secFGF in which AATPAP was introduced at the N- or C-terminus of secFGF were constructed in a eukaryotic expression vector. [AATPAP]secFGF proteins were then expressed in Chinese hamster ovary (CHO) cells and secreted into the surrounding medium, primarily as modified forms sensitive to sialidase but not to peptide N-glycosidase F. The modifying groups were not seen when the AATPAP sequence was converted to AAAPAP or when [AATPAP]secFGF was expressed in mutant cells incapable of UDP-GalNAc biosynthesis. The results indicate that the modifying groups were mucin-type O-glycans and that the AATPAP served as an efficient O-glycosylation signal sequence. The O-glycosylated forms of [AATPAP]secFGF were as mitogenic toward human vascular endothelial cells as unmodified secFGF, suggesting that introduction of the signal into biologically active polypeptides is a promising approach with which O-glycosylation may be achieved without affecting original activity.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1007092708666
