Title : The structural role of N-linked glycans on human
glypican-1
Abstract :
- Glypicans are cell-surface heparan sulfate proteoglycans that regulate developmental signaling pathways by binding growth factors to their heparan sulfate chains
- The primary structures of glypican core proteins contain potential N-glycosylation sites , but the importance of N-glycosylation in glypicans has never been investigated in detail
- Here, we studied the role of the possible N-glycosylation sites at Asn-79 and Asn-116 in recombinant anchorless glypican-1 expressed in eukaryotic cells
- Mutagenesis and enzymatic cleavage indicated that the potential N-glycosylation sites are invariably occupied
- Experiments using the drug tunicamycin to inhibit the N-linked glycosylation of glypican-1 showed that secretion of anchorless glypican-1 was reduced and that the protein did not accumulate inside the cells
- Heparan sulfate substitution of N-glycosylation mutant N116Q was similar to wild-type glypican-1 while the N79Q mutant and also the double mutant N79Q,N116Q were mostly secreted as high-molecular-weight heparan sulfate proteoglycan
- N-Glycosylation mutants and N-deglycosylated glypican-1 had far-UV circular dichroism and fluorescence emission spectra that were highly similar to those of N-glycosylated glypican-1
- A single unfolding transition at high concentrations of urea was found for both N-deglycosylated glypican-1 and glypican-1 in which the N-glycosylation sites had been removed by mutagenesis when chemical denaturation was monitored by circular dichroism and fluorescence emission spectroscopy
- In summary, we have found that the potential N-glycosylation sites in glypican-1 are invariably occupied and that the N-linked glycans on glypican-1 affect protein expression and heparan sulfate substitution but that correct folding can be obtained in the absence of N-linked glycans
Output (sent_index, trigger,
protein,
sugar,
site):
- 2. N-glycosylation, , -, -, sites
- 3. N-glycosylation, , -, -, Asn-79 and Asn-116
- 3. N-glycosylation, , -, -, sites
- 4. N-glycosylation, , -, -, sites
- 4. occupied, , -, -, sites
- 5. glycosylation, , glypican-1, -, -
- 7. N-deglycosylated, , N-deglycosylated glypican-1, -, -
- 7. N-glycosylated, , N-glycosylated glypican-1, -, -
- 8. N-deglycosylated, , N-deglycosylated glypican-1, -, -
- 8. N-glycosylation, , -, -, sites
- 9. N-glycosylation, , -, -, sites
- 9. glycans, , glypican-1, glycans, -
- 9. occupied, , -, -, sites
Output(Part-Of) (sent_index,
protein,
site):
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 3. glypican-1, -, Asn-79 and Asn-116